Journal of

Clinical Medicine

Review

Biodistribution of Mesenchymal Stromal Cells after

Administration in Animal Models and Humans:

A Systematic Review

Manuel Sanchez-Diaz ¹

, Maria I. Quiñones-Vico ^2,*, Raquel Sanabria de la Torre ², Trinidad Montero-Vílchez ¹

Alvaro Sierra-Sánchez ²

, Alejandro Molina-Leyva ¹

and Salvador Arias-Santiago ^1,2,3

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Citation: Sanchez-Diaz, M.;

Quiñones-Vico, M.I.; Sanabria de la

Torre, R.; Montero-Vílchez, T.;

Sierra-Sánchez, A.; Molina-Leyva, A.;

Arias-Santiago, S. Biodistribution of

Mesenchymal Stromal Cells after

Administration in Animal Models

and Humans: A Systematic Review. J.

Clin. Med. 2021, 10, 2925. https://

doi.org/10.3390/jcm10132925

Academic Editor: Kyung-Rok Yu

Received: 12 May 2021

Accepted: 25 June 2021

Published: 29 June 2021

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Attribution (CC BY) license (https://

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4.0/).

Dermatology Department, Hospital Universitario Virgen de las Nieves, IBS Granada, 18014 Granada, Spain;

manolo.94.sanchez@gmail.com (M.S.-D.); tmonterov@gmail.com (T.M.-V.);

alejandromolinaleyva@gmail.com (A.M.-L.); salvadorarias@ugr.es (S.A.-S.)

Cellular Production Unit, Hospital Universitario Virgen de las Nieves, IBS Granada, 18014 Granada, Spain;

raquelsanabriadlt@gmail.com (R.S.d.l.T.); alvarosisan@gmail.com (A.S.-S.)

School of Medicine, University of Granada, 18014 Granada, Spain

Correspondence: maribelmqv20@gmail.com

Abstract: Mesenchymal Stromal Cells (MSCs) are of great interest in cellular therapy. Different routes

of administration of MSCs have been described both in pre-clinical and clinical reports. Knowledge

about the fate of the administered cells is critical for developing MSC-based therapies. The aim of this

review is to describe how MSCs are distributed after injection, using different administration routes

in animal models and humans. A literature search was performed in order to consider how MSCs

distribute after intravenous, intraarterial, intramuscular, intraarticular and intralesional injection

into both animal models and humans. Studies addressing the biodistribution of MSCs in “in vivo”

animal models and humans were included. After the search, 109 articles were included in the review.

Intravenous administration of MSCs is widely used; it leads to an initial accumulation of cells in the

lungs with later redistribution to the liver, spleen and kidneys. Intraarterial infusion bypasses the

lungs, so MSCs distribute widely throughout the rest of the body. Intramuscular, intraarticular and

intradermal administration lack systemic biodistribution. Injection into various speciﬁc organs is also

described. Biodistribution of MSCs in animal models and humans appears to be similar and depends

on the route of administration. More studies with standardized protocols of MSC administration

could be useful in order to make results homogeneous and more comparable.

Keywords: mesenchymal stromal cell; biodistribution; cell therapy

1. Introduction

Mesenchymal Stromal Cells (MSCs) are non-hematopoietic multipotent cells which can

be isolated from different tissues from adult, perinatal and fetal samples [1,2]. Some sources

are adipose tissue [3], bone marrow [4], umbilical cord Wharton’s jelly and blood [5,6],

periosteum [7], skin [8], amniotic ﬂuid [9] and the placenta [10]. These cells have the

capability to differentiate into a variety of different mesenchymal lineage cells such as

osteoblasts, chondrocytes, adipocytes, ﬁbroblasts and myoblasts [2].

Since MSCs have variable phenotypes, with different expression of bio-markers de-

pending on the source and means of isolation, as well as the tissue they come from, they

cannot be considered as a homogeneous set of cells [11]. The International Society for

Cellular Therapy set minimum criteria for characterizing human MSCs in order to pro-

mote a more uniform deﬁnition of MSCs. These criteria are: (a) Plastic adherence when

maintained in standard culture conditions; (b) Expression of CD105, CD73 and CD90

and lack of expression of CD45, CD34, CD14, or CD11b, CD79a or CD19 and HLA-DR

surface molecules; and (c) Differentiation into osteoblasts, adipocytes and chondroblasts

in vitro [12].

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MSCs are of great interest because of the possibility of using them as a part of ther-

apeutic regimens in a wide variety of human diseases, e.g., rheumatic and autoimmune

diseases, skin diseases and complex ulcers and wounds [13–16]. Some characteristics of

MSCs are fundamental for this purpose: (a) MSCs can be obtained from adult donors and

expanded in vitro without losing their immunomodulatory and differentiation potential;

(b) MSCs have hypo-immunogenic properties, so allogenic sets can be used, avoiding the

need for autologous cell cultures; (c) Their immunomodulatory and transdifferentiating

capabilities into different cell lineages can be exploited as a novel approach to the treatment

of different diseases [13–15,17–19].

Different routes of administration of MSC-based medical therapies have been de-

scribed both in pre-clinical and clinical reports, and the possible differences between them,

in terms of safety and efﬁcacy, is an issue which is still under discussion [15,16,20–23].

These differences may be explained by the variable biodistribution of MSCs after their ad-

ministration. The most common reported routes of administration are topical, intravenous

and intraarterial, intramuscular and intralesional (including different locations e.g., skin,

spinal cord, tendons).

Given the presumable importance of the different mechanisms of MSC biodistribution

and their impact on the therapeutic effects, the objective of this systematic review is to

describe how MSCs are distributed after their inoculation through different administration

routes in animal models and humans.

2. Materials and Methods

2.1. Search Strategy

A literature search from January 2015 to April 2021 was performed using the Medline

database. The following search terms were used: MSC or MESENCHYMAL STEM CELL

or MESENCHYMAL STROMAL CELL or MULTIPOTENT STEM CELL or MULTIPOTENT

STROMAL CELL or STEM CELL AND BIODISTRIBUTION or DISTRIBUTION.

2.2. Inclusion and Exclusion Criteria

The search was limited to: (a) Human or animal data; (b) In vivo studies; (c) Studies

addressing the biodistribution of MSCs after any source of administration; (d) Articles

written in English or Spanish. All types of epidemiological studies (clinical trials, cohort

studies, case-control studies and cross-sectional studies) regarding the biodistribution of

MSCs were considered.

2.3. Study Selection

The titles and abstracts obtained in the ﬁrst search were reviewed to assess relevant

studies. The full texts of all articles meeting the inclusion criteria were reviewed and their

bibliographic references were checked for additional sources. Articles considered relevant

were included in the analysis. Uncertainties about the inclusion or exclusion of articles

were subjected to discussion until a consensus was reached.

2.4. Research Questions and Variables Assessed

The research questions were as follows:

•

How do MSCs distribute after intravenous and intraarterial injection in animal models

and humans?

•

How do MSCs distribute after intramuscular injection in animal models and humans?

•

How do MSCs distribute after intralesional injection in different organs and tissues in

animal models and humans?

•

Which cell marking techniques have recently been used in studies on humans?

The variables assessed in order to answer these questions were the model which

received the MSCs (human or animal), the route of administration, the disease treated, the

cell-marking technique used, the biodistribution assessment method, the time when the

assessment was performed, and the outcomes regarding the biodistribution of the MSCs.

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3. Results

An initial search found 6808 references (see Figure 1). After reviewing the titles and

abstracts, 159 articles underwent full-text review. From this list, 50 articles were eventually

discarded due to various issues: 33 articles did not assess biodistribution; 7 were related

to other types of cells, rather than MSCs; 6 were not accessible or written in a different

language; 3 only addressed the issue of in vitro MSCs; and 1 article was duplicated. Finally,

109 studies met the eligible criteria and were included in the review.

Figure 1. Search strategy.

3.1. Biodistribution Characteristics of Mscs Depending on the Route of Administration

An overview and summary of all the information collected in this study can be seen

in Table 1.

3.1.1. MSC Biodistribution in Animal Models

First, the biodistribution of MSCs after their delivery or injection into animal models

will be discussed. Intravenous and intraarterial infusion, intramuscular injection and a

wide variety of intralesional administrations of MSCs will be addressed in this section.

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Table 1. Overview of the characteristics of each route of administration.

Route of

Administration

Systemic

Distribution

Organs to Which the Cells are

Distributed

Advantages

Disadvantages

Intravenous

Yes

First, cells move to the lungs

(ﬁrst capillary ﬁlter).

Later, cells distribute, mainly to

the liver, spleen and kidneys.

Variable amounts of cells are

found in other organs.

Convenient route of

administration.

Widely used.

Useful to reach the

lungs.

Cells do not reach other

organs apart from

lungs in great

quantities.

Intraarterial (not

selective)

Yes

Cells bypass the pulmonary

ﬁlter so there is a wide

distribution in the rest of the

organs (heart, brain, kidneys,

liver, digestive system)

Convenient route of

administration.

Useful to bypass the

lungs and achieve

broader distribution.

Not so widely used.

Intraarterial infusion is

not common in clinical

practice

Intraarterial

(selective)

Yes (reduced)

Cells are distributed mainly in

the territory irrigated by the

cannulated artery. Distribution

of cells to other organs is

possible but in smaller amounts.

Targeted deposition of

cells is achieved.

Inconvenient route of

administration.

Difﬁcult to transfer to

clinical practice

Intramuscular and

intraarticular

Cells remain at the injection site

Convenient route of

administration.

Targeted deposition of

cells is achieved

No systemic

distribution is

achieved.

Intradermal,

intratracheal,

intrapulmonary

and intraurinary

tissue

Cells remain at the injection site

Convenient route of

administration,

depending on each

speciﬁc route.

Targeted deposition of

cells is achieved

No systemic

distribution is

achieved.

Intrahepatic,

intrasplenic,

intrapericardial,

intramyocardial

Yes

Cells distribute following the

direction of the bloodstream

derived from the infused organ.

Targeted deposition is

achieved.

Knowledge about the

bloodstream derived

from the infused organ

might lead to targeted

distribution after

injection.

Inconvenient in clinical

practice.

Difﬁcult to transfer to

clinical practice.

Injection into

cavities containing

body ﬂuids

(peritoneum,

cerebral ventricles)

Yes (low

amounts)

Cells distribute mainly to tissues

in contact with the body ﬂuid.

Convenient route of

administration,

depending on each

speciﬁc route.

Targeted deposition is

achieved.

Limited systemic

biodistribution.

Limited systemic

biodistribution.

Intrathecal

administration

Cells distribute caudally when

injected in the upper segments of

spine. Cranial migration of cells

after lumbar injection seems to

be possible if a high dose of

MSCs is administered (e.g.,

distribution to brain).

Convenient route of

administration if

deposition of the cells

at the central nervous

system level is desired.

Limited systemic

biodistribution.

Inconvenient in clinical

practice (depending on

the cases).

Intra-Central

Nervous System

Yes/No (variable

amounts)

Cells are able to distribute within

the central nervous system.

Factors leading to the movement

of the cells are still not clear.

Targeted deposition is

achieved.

Inconvenient route of

administration.

Difﬁcult to transfer to

clinical practice.

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3.1.2. Distribution of MSCs after Intravenous Injection in Animal Models

Intravenous injection has emerged as the most widely used route in the various

research studies. This route of administration is a simple and effective way to deliver

MSCs systemically. Most of the studies discussed in this section agree on the general

characteristics of how mesenchymal cells are distributed after being injected into the

venous stream (Table 2, Figure 2). To begin with, some general ideas can be stated about

this issue: (a) after IV injection, most cells are retained initially in the lungs, which is

the ﬁrst capillary ﬁlter; (b) there is later redistribution of the cells, mainly to the liver,

spleen and kidney, with few MSCs redistributing to other organs; (c) in some studies,

later redistribution is very limited; and (d) some pathological entities seem to alter this

biodistribution pattern.

A good example of this general distribution pattern can be seen in one study assessing

intravenous infusion of MSCs in a myocardial infarction model in dogs [24]. It showed high

distribution during the immediate post-infusion time in the lungs, with a posterior decrease

in the amount of MSCs and a later redistribution from day 1 to 7 in different tissues, mainly

in the liver, spleen and kidney. A similar model of myocardial infarction in mice [25]

showed early distribution in lungs but an insigniﬁcant amount of cells distributed to other

organs (less than 1%). Intravenous infusion of MSCs in baboons [26], and a late evaluation

of their distribution in a variety of tissues, have demonstrated a wide distribution of

MSCs after a long period of time: gastrointestinal, kidney, skin, lung, thymus, and liver

tissues contained MSCs. Similar results were shown in several other studies [27–31]. The

redistribution might be explained by phagocitation of MSCs: monocytes might perform

this action, and then change their immunophenotype, inducing Treg cells [32].

Figure 2. Biodistribution of MSCs after intravenous infusion. After intravenous infusion, there is initial biodistribution in

the lungs. Later, most cells redistribute to the liver, kidney and spleen. Few cells can be found in other organs and tissues.

In some cases, diseased tissues have been found to be capable of attracting MSCs.

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The alteration of the general distribution pattern in speciﬁc diseases has also been

reported in several studies. Zhang et al. [33] found a signiﬁcant amount of MSCs in the

kidneys of rabbits with acute kidney injury. Similar results have been shown in a model of

Alzheimer’s disease [34,35] with higher brain distribution of MSCs in diseased animals

compared to healthy animals. This was also evidenced in another study performed on

mice with cerebral tumors [36], rats and beagle dogs with spinal cord injury [37,38], and

rats with intracerebral hemorrhage [39]. Moreover, acute distress respiratory syndrome or

liver tumors may also affect the distribution of cells after intravenous injection [40,41]. In

contrast, in a murine model of experimental autoimmune encephalomyelitis [42], MSCs

were not distributed to the brain area.

Although the lungs seem to be the area MSCs mostly distribute to after intravenous

injection, Schmuck et al. [43] concluded that this may be due to the lack of sensitivity of

bioluminescence techniques, which are carried out in most biodistribution studies. In their

study, which used a 3D cryo-imaging system, they demonstrated a higher concentration of

MSCs in the liver when compared to the lungs after intravenous infusion in rats with acute

lung injury. In this line, Schubert et al. [44] demonstrated a high distribution of MSCs to

the lungs with bioluminescence techniques on day 1 after intravenous infusion in mice

with acute kidney injury. Cells cleared on days 3 and 6. However, when RT-PCR was

performed on several tissues on day 6, variable amounts of mRNA were detected in the

blood, liver, kidneys and lungs. Therefore, RT-PCR could be a better option for detecting

the late presence of MSCs in tissues and could be used to complement imaging techniques.

Other situations, such as the modiﬁcation of MSCs or the selective infusion of MSCs

into certain veins, might also affect biodistribution. Moreover, some studies have shown

that modifying MSCs may lead to cells selectively targeting speciﬁc organs. The modiﬁ-

cation of speciﬁc “homing markers” or adhesion molecules can lead to targeted homing

of MSCs. This has been proven by modifying MSCs to achieve speciﬁc distribution to

the liver [30]. In addition, the selective intravenous delivery could lead to differences in

biodistribution. For example, Li et al. [45] demonstrated that superior mesenteric vein

infusion of MSCs leads to more selective and longer homing of MSCs in a model of acute

liver injury when compared to intravenous and inferior vena cava delivery.

Finally, regardless of the source of administration, Fabian et al. [46] demonstrated that

the age of both the recipient and the donor of MSCs seems to affect the biodistribution of

the cells. The study demonstrated that old recipients and donors showed a very restricted

biodistribution of MSCs in mice after 28 days (mainly in the brain cortex and spleen)

whereas young receptors and donors showed a wide variety of distribution.

3.1.3. Distribution of MSCs after Intraarterial Injection in Animal Models

Intraarterial infusion of MSCs has been used as an alternative and has also been

compared to IV injection in several situations (Table 3, Figure 3). Brieﬂy, the main charac-

teristics of this route of administration are: (a) IA injection bypasses the pulmonary ﬁlter,

so low amounts of MSCs are retained in these organs; (b) MSCs distribute more widely

into the rest of the body’s organs after IA infusion compared to IV delivery; (c) like the IV

route, biodistribution after IA injection might be modiﬁed by several diseases; d) selective

intraarterial delivery of MSCs might be very useful for targeting diseased organs.

As an initial example of these characteristics, one study performed on pigs [22]

compared intravenous and intraarterial infusion techniques. MSCs were detected using

SPECT/TC imaging, which showed a lower pulmonary captation in the intraarterial group,

and a relatively higher uptake in other organs such as the liver, spleen and kidney. This

was also studied in an acute kidney injury model in mice [47]. In this study, a signiﬁcantly

higher amount of MSCs were detected in the kidneys after intraarterial infusion, especially

in mice with AKI. In contrast, the vast majority of MSCs were distributed to the lungs

after intravenous injection. Moreover, intracardiac injection has also been reported to be an

effective delivery route. This route of administration can be considered to be equivalent

to the IA route when the cells are injected into the left chambers of the heart. In fact, after

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intracardiac injection [48], MSCs seem to follow a similar path; widespread distribution is

observed (lungs, brain, spleen, liver, kidneys).

The fact that the IA route leads to a signiﬁcantly higher distribution of MSCs in

peripheral organs might be an interesting characteristic when homing MSCs in the diseased

area is desirable. For example, in other studies it has been proven that intraarterial injection

improves distribution to the damaged cerebral areas when compared to intravenous

injection [49–51].

Regarding the distribution of MSCs to the brain after intraarterial infusion, Cerri

et al. [52] evaluated distribution to the brain of MSCs injected in the carotid artery of a

Parkinson’s disease murine model. One group was treated with mannitol as a transient

permeabilizing factor of the blood-brain barrier. Later assessment showed that rats not

treated with mannitol had an extremely low amount of MSCs homing to the brain, whereas

the group treated with mannitol showed a signiﬁcantly higher amount of MSCs. Moreover,

most of the cells were distributed in the ipsilateral hemisphere to the carotid used to inject

them. Therefore, the use of a permeabilizing agent could be essential to allow the passage

of MSCs into the brain. On the other hand, selective delivery of cells might help MSCs

reach the damaged areas [51,53].

Figure 3. Biodistribution of MSCs after intraarterial infusion. When cells are administered into a peripheral artery, the lungs

are bypassed and a wide distribution of cells is found in organs and tissues. Selective intraarterial delivery of cells targets

the distribution of cells to organs which are irrigated by the cannulated artery.

As occurred with IV injection, some pathological entities can modify the biodistri-

bution of MSCs after IA injection. In the speciﬁc case of mice with inﬂammatory bowel

disease, MSCs do not signiﬁcantly distribute to lungs or liver but distribute mainly to

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the affected areas of the intestine [54]. In contrast, in a model of kidney injury, MSCs did

not distribute to damaged kidneys after intracardiac injection [55]. Moreover, the dose of

MSCs seems to be important when administered IA. One study showed that an increased

dose of IA-administered MSCs led to a wider distribution of cells but also to a high degree

of intravascular cell aggregation and mortality [56]. Thus, the dose of MSCs should be

assessed before intraarterial delivery to avoid intraarterial aggregation.

The homing of MSCs to diseased tissues can be improved by selective intraarterial

infusion. With this technique, MSCs are directly injected into selected arteries. This results

in a greater amount of MSCs in the targeted organs. Some examples are discussed here:

When MSCs are delivered directly into the renal artery, MSCs seem to distribute only in

the kidneys, without systemic signiﬁcant distribution, and mainly in the renal cortex [57].

Therefore, renal intraarterial MSC infusion limits off-target engraftment, leading to efﬁcient

MSC delivery to the kidneys. Similar results were found after selective intraarterial infusion

into the superior mesenteric artery regarding the intestine distribution of MSCs [58], and

the selective intraarterial limb infusion [59,60], with MSCs distributed in the target area

and a small quantity of MSCs in the rest of the organs.

3.1.4. Distribution of MSCs after Intramuscular Injection in Animal Models

As this is widely used with classic drugs, intramuscular injection of MSCs has also

been studied as a possible way to administrate MSCs (Table 4, Figure 4). As a general

idea, whereas intramuscular injection of conventional drugs leads to a signiﬁcant systemic

distribution, MSCs injected intramuscularly do not seem to distribute to the rest of the body.

One study performed on mice to assess the sensitivity and speciﬁcity of quantitative

PCR [61] for detecting MSCs showed that, 3 months after intramuscular injection of MSCs,

no MSCs were detectable in any internal organ. However, DNA from MSCs was still present

in the muscles where it was injected. This could suggest that MSCs do not distribute to other

organs after intramuscular injection. This was in line with the ﬁndings of similar studies

performed following intramuscular injection [62–64], with MSCs remaining at the injection

site, but without MSCs distributing to organs. However, it has been demonstrated that,

despite the lack of distribution of MSCs, when injected intramuscularly in a contralateral

muscle to an inﬂamed area, MSCs are capable of reducing inﬂammation. This is thought to

be performed by the release of soluble factors rather than the movement of the cells [65].

A recent review of intramuscular MSCs showed that, to date, no articles have found

signiﬁcant systemic biodistribution after intramuscular injection of MSCs [65].

3.1.5. Distribution of MSCs after Intralesional Injection in Animal Models

Several different intralesional routes of administration for MSC delivery have been

described. The most important routes of administration of MSCs into lesioned areas will

now be addressed

•

Intraarticular (IAr) delivery of MSCs (Table 4, Figure 4):

Intraarticular injection of MSCs has been widely studied in different animal models.

As a general idea, IAr injection lacks systemic biodistribution, whereas it leads to a very

targeted delivery of cells into the joints. This has been adequately demonstrated by studies

on different mice models of healthy animals, arthritis and osteoarthritis, where it was shown

that MSCs do not distribute to other organs following intraarticular injection [21,66–69].

Markides et al. [70] assessed the biodistribution of MSCs in a sheep model of osteochondral

injury. After intraarticular injection, MSCs were only detected in the synovium, with a

lack of MSCs within the chondral defect. Khan et al. [71] showed similar results after

intratendinous injection, with no MSCs spreading from the injection site.

In contrast with that already described, some studies show an incidental distribution

of MSCs. In these cases, MSCs have been shown to be present in the blood, distant zones

or tendon lesions near the injection site. One study performed on a horse model of tendon

lesions [72] showed that, although the vast majority of cells remained at the site where they

had been injected, a small amount of MSCs could be found in blood for the ﬁrst 24 h after

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injection, as well as in the contralateral control tendon lesions which had not been injected.

Similar results were observed by Shim et al. [73]; after intraarticular injection, MSCs were

detectable in blood with a peak at 8 h. No systemic distribution was observed. Moreover,

other studies show that MSCs seem to be able to migrate from the joint to nearby tendinous

lesions [74,75].

Figure 4. Biodistribution of MSCs after intraarticular and intramuscular injection. No systemic distribution has been

demonstrated after intramuscular or intraarticular injection. After intraarticular injection, MSCs have been found to be able

to migrate to nearby damaged lesions and into the bloodstream. Moreover, the use of a magnet on MSCs with a magnetic

label is useful for targeted deposition of cells within the joint.

As occurred with the IV and IA routes, elective accumulation of MSCs in selected areas

of a joint (i.e., a chondral lesion within the joint) can be achieved. MSCs must be modiﬁed

by magnetic labeling. The subsequent use of a magnet during the transplantation [76] leads

to the movement of the cells within the joint so they can be deposited in the target zone.

Finally, as a variant of IAr delivery, one study was performed to assess biodistribution

of MSCs which were pre-loaded into bone grafts [77]. This study also showed the lack of

systemic biodistribution of MSCs and the long-lasting MSCs in the graft up to 6 weeks.

Similar results were found when injecting MSCs into the femoral head of pigs [78].

3.1.6. Injection of MSCs into the Reproductive and Urinary System

Some studies have been found on the issue of biodistribution of MSCs after injection

into the urinary and reproductive systems. In a rat model of birth-trauma injury [79], the

presence of MSCs following local injection into the periurethral tissues was demonstrated

up to 7 days post-injection. In this case, no tests were performed to assess the distribution

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to other organs after local injection. Ryu et al. [80] injected MSCs into the outer layer of the

bladder in a interstitial cystitis model. It was demonstrated that cells are able to migrate

from the outer layers of the bladder to the urothelium for the ﬁrst 30 days after injection

and to home as perivascular cells. Dou et al. [81] found that after intracavernous injection,

MSCs distributed to the lower abdomen in a erectile dysfunction model in mice in the ﬁrst

hour. Moreover, MSCs can be found in kidney, prostate and hepatic tissues up to 7 days

after injection. Finally, when injected into the ovaries, MSCs are able to distribute to the

uterus, with no systemic distribution Table 5 [82].

3.1.7. Injection of MSCs into the Central Nervous System

There is a wide variety of reports concerning the injection of MSCs into the central

nervous system Table 6: Intrathecal, intracerebral and intraventricular injections have been

described:

(a)

Intrathecal injection of MSCs: After intrathecal injection, Barberini et al. [83] demon-

strated that MSCs do not seem to distribute cranially (when injected in the lumbosacral

area), whereas they can progress caudally (when injected in the altanto-occipital area).

In this study, no MSC engraftment was demonstrated. The systemic biodistribution

of the MSCs was not speciﬁcally assessed, but imaging techniques did not show

the presence of MSCs in areas other than the central nervous system. In contrast,

Kim et al. [84] demonstrated that MSC migration from the spine to the brain is pos-

sible in a dose-dependent manner. Quesada et al. [85] also demonstrated brain

migration after intrathecal injection;

(b)

Intracerebral injection of MSCs: Wang et al. [86] demonstrated that intracerebrally

injected MSCs loaded with paclitaxel are capable of spreading from one cerebral

hemisphere to another in a glioma model in mice in two days. These cells were found

to spread from the healthy hemisphere to the glioma hemisphere and to invade the

tumor. The ability of MSCs to migrate from one hemisphere to another has also

been demonstrated in other studies [87]. In other reports [88–93], MSCs injected

intracerebrally were detectable at the site of administration 1–3 weeks after injection,

with a subsequent rapid decrease and no signiﬁcant systemic distribution. Other

studies [94] showed that MSCs can be detected with ﬂuorescence and bioluminescence

up to 7 weeks after transplantation;

(c)

Intraventricular injection of MSCs: Some studies showed that MSCs injected into

cerebral ventricles are able to migrate to large blood vessels in a brain traumatic injury

model [95], and also to brain parenchyma and the spinal cord [96]. In contrast, other

reports [97] demonstrate that after intraventricular infusion, MSCs do not migrate

to brain parenchyma and are hardly able to migrate to the spinal cord in a model of

amyotrophic lateral sclerosis.

Finally, one review showed that intranasal delivery of MSCs led to signiﬁcant intrac-

erebral migration of MSCs [98].

3.1.8. Injection of MSCs into the Digestive System:

(a)

Intrahepatic and intrasplenic injections have been studied in several reports as efﬁcient

delivery routes for administrating MSCs. After intrahepatic injection, Xie et al. [99]

demonstrated that MSCs remain in the liver and are cleared in a short period of time,

without systemic distribution. This short period of time might be in association with

NK cell activation: Liu et al. [100] showed that mice with activated NK cells had a

more rapid clearance of intrahepatic MSCs. Yaochite et al. [101] injected MSCs into

the liver and spleen of diabetic mice. It was shown that intrasplenic MSCs were able

to move to the liver whereas intrapancreatic cells remained at the site of the injection.

No systemic distribution was shown and cells remained for up to 8 days. Similar

results were found in another study [102], with MSCs remaining for up to 4 weeks;

(b)

When injected intraperitoneally [103,104], MSCs seem to spread mostly to abdominal

organs (liver, spleen and intestine) with little distribution to the lungs, heart, blood

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and lymph nodes. Other study shows that Wharton’s Jelly MSCs are capable of

distributing to the whole body after intraperitoneal injection at days 1, 7, 14 and 21 in

piglets [105].

(c)

When injected in the peri-fistula area [106], MSCs do not seem to distribute systemically.

3.1.9. Injection of MSCs into the Cardiovascular and Respiratory Systems

Some articles have addressed the injection of MSCs into the pericardium or the my-

ocardium. When injected intrapericardially [107] in a myocardial infarction model, MSCs

seem to distribute to ventricles and atriums, with a preference for the left ventricle. Regard-

ing intramyocardial injection, MSCs seem to distribute initially in the myocardium, with

posterior redistribution to the lungs, liver and bone [108]. Moreover, it has been demon-

strated that after intramyocardial injection, if a repeated ischemia model is performed,

MSCs tend to home mainly to the heart with less distribution to peripheral organs [109]. Fi-

nally, some studies were related to the injection of MSCs into the respiratory system: When

injected intratracheally or intrabronchially, MSCs do not distribute systemically [110,111].

3.1.10. Injection into the Skin, Subcutaneous Cellular Tissues and Lymph Nodes

Few studies have addressed the issue of biodistribution after intradermal injection of

cells (Table 5). When injected into the skin of mice, Tappenbeck et al. [112] demonstrated

that MSCs seem to remain in the skin and migrate to lymph nodes, without signiﬁcant

systemic distribution. Regarding the speciﬁc distribution in cutaneous wounds [113,114],

MSCs seem to distribute with a diffuse pattern initially and later concentrate towards the

wound edges. Finally, these cells seem to be engrafted with the newly developed skin

tissue. No systemic distribution following intradermal injection had been reported. Only

one study was performed to describe biodistribution after intranodal injection; in this

study, most MSCs remain at the injection site or in the fat surrounding the injected nodes

48 h later [103], without systemic distribution of cells.

3.2. Biodistribution of MSCs in Humans

Only a few reports of the biodistribution of MSCs after the injection into human

models have been recorded in this review (Table 7). These articles will be discussed in the

following sections.

3.2.1. Distribution of MSCs after Intravenous Injection in Humans

Three studies regarding the intravenous injection of MSCs into humans were identiﬁed

in order to assess biodistribution. In the ﬁrst study, the intravenous infusion of MSCs in

patients suffering from cirrhosis showed an early (pre-48 h) distribution mainly in the

lungs, with a later decrease in lung captation and a high distribution in the spleen and

liver [115]. In another study on breast cancer patients, MSCs were monitored in peripheral

blood after intravenous injection [116], ﬁnding a rapid clearance of MSCs in blood, with

no cells detected 1 h post injection. Finally, a third article showed that when injected

intravenously into a patient with hemophilia A [117], MSCs distributed early to the lungs

and liver, with a progressive decrease. Distribution to the usual bleeding places was shown

at 24 h.

As can be seen, biodistribution after IV injection in humans seems to be similar to

that described in animal models: (a) early captation in the lungs; (b) later distribution in

organs such as the spleen and liver; and (c) distribution of MSCs into target areas have

been described.

3.2.2. Distribution of MSCs after Intraarterial Injection in Humans

Only one study addressed the intraarterial infusion of MSCs in humans. This study

was performed on 21 type 2 diabetes mellitus patients. MSCs were selectively injected

intravenously or intraarterially (into the pancreaticoduodenal artery and the splenic artery).

MSCs were labeled with 18-FDG and PET-TC images were used to assess biodistribution.

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Selective intraarterial delivery led to MSCs homing to the pancreas head (when cells

were injected into the pancreaticoduodenal artery) or body (when infused into the splenic

artery); whereas no MSCs were found in the pancreas in the intravenous group. This report

shows that biodistribution after IA infusion of MSCs seem to be similar to biodistribution

in animal models, with systemic delivery, a lack of lung trapping and the possibility of

selective infusion into certain areas.

3.2.3. Distribution of MSCs after Intralesional Injection in Humans

Only one study of intralesional injection of MSCs and their biodistribution was ob-

served. Henriksson et al. [118] injected MSCs into intervertebral discs in 4 patients. Discs

were explanted at 8 and 28 months post injection. Histologic examinations found the pres-

ence of MSCs in intervertebral discs after 8 months, with chondrocyte-like differentiation.

No cells were found in the discs after 28 months, and no systemic distribution was assessed.

3.2.4. Distribution of MSCs after Intracoronary Injection in Humans

In one study, biodistribution of MSCs after intracoronary injection was assessed.

Lezaic et al. [119] injected MSCs into the coronary arteries of 35 patients with idiopathic di-

lated cardiomyopathy. They showed that a very low number (0–1.25%) of MSCs are retained

in the myocardium, with the majority distributed to the liver, spleen and bone marrow.

3.2.5. Which Cell-Marking Techniques Have Recently Been Used in Preclinical Studies?

A wide variety of cell-marking techniques have been used for preclinical studies

involving cell therapy. Also, a wide variety of detection methods have been performed.

Since it is not the objective of this review to address these techniques in depth, an overview

of them is reviewed hereafter.

Most common cell-marking techniques can be divided into: (a) those related to the use

of radionuclides; (b) those related to bioluminescence imaging systems; (c) those related to

the use of magnetic resonance imaging (MRI); and (d) those related to the genetic marking

of cells.

The use of radionuclides is a common technique which is useful to assess the distribu-

tion of previously marked cells in preclinical models. Some of the most common radionu-

clides include ^99mTechnetium-hexamethylpropyleneamine oxime (^9mTc-HMPAO) [21,83],

or ¹¹¹Indium-Oxine (¹¹¹In-Oxine) [60,115]. After culture, these substances enter into the

cells. Once the cells are administered, the emitted radioactivity can be detected by imaging

techniques such as Single Photon Emission Computed Tomography (SPECT) or Positron

Emission Tomography (PET), which allow us to track the fate of the cells. The main dis-

advantage of these methods are the limited duration of the radioactivity, which limits the

assessment of the late distribution of cells, and the dangers related to the management of

radioactive substances in the laboratory.

Bioluminescence imaging systems are based on the light which is emitted by cells

which have been previously transfected with the ﬁreﬂy luciferase gene (luc gene) [99,100].

Once the cells transfected with this gene are administered to the animal, an injection of D-

luciferin is performed. After D-luciferin has been administered, cells containing the speciﬁc

gene ﬂuoresce, emit light with a wavelength of 537 nm. This light can be detected by

speciﬁc imaging systems to assess biodistribution. The main disadvantage of this method

is that bioluminescence has limited spatial resolution and reduced tissue penetration due

to the relatively weak energy of emitted photons. For these reasons, its use in large animal

models is not advisable [120].

The use of superparamagnetic iron oxide nanoparticles (SPIONs) is also a useful

technique to assess cell biodistribution. SPIONs are small synthetic iron particles which

are coated with certain biocompatible polymers. When cells have been labeled with these

particles, they are detectable by imaging techniques such as MRI techniques [121]. Given

that magnetic resonance imaging is a technique that is not very accessible, the use of this

method can be limited.

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Finally, it is possible to label cells with speciﬁc genes that can be subsequently detected

by PCR methods [26,44,105]. The main disadvantage of this cell-marking technique is that

a tissue sample is required so that the distribution of cells cannot be assessed in vivo in

most cases.

3.2.6. Which Cell-Marking Techniques Have Recently Been Used in Studies with Humans?

The studies included in this review used different cell-marking techniques. The most

common techniques are those related to the use of a radioactive labeling: MSCs can be la-

beled with radionuclides in vitro, and then injected into humans. Radionuclides used in the

reviewed articles include ¹¹¹Indium-Oxine (¹¹¹In-Oxine) [115,117], 18-Fluorodeoxyglucose

(¹⁸F-FDG) [122] and ^99mTechnetium-hexamethylpropyleneamine oxime (^9mTc-HMPAO) [119].

Cells are incubated in culture mediums containing these substances, which enter into the

cells. Later assessment of the emitted radioactivity of these substances in the body with

imaging techniques such as scintigraphy, Single Photon Emission Computed Tomogra-

phy (SPECT) or Positron Emission Tomography (PET) allow us to detect the distribution

of the cells in the body to be evaluated. The main disadvantage of radionuclide-based

cell-marking techniques is the limited duration of the radioactivity; as the radionuclides dis-

integrate, the emitted signal becomes smaller and ﬁnally disappears, making it impossible

to evaluate the late biodistribution of the cells administered.

Labeling cells with markers which can be detected in histologic samples is another

technique used in humans. In the study reviewed [118], iron sucrose was used to label

cells. This compound makes cells detectable in histologic samples. The main advantage of

this kind of marker is its presumably long duration (longer than radionuclides). However,

the use of histologic markers makes it necessary to perform ex vivo examination which

is a limiting factor for its use in humans. The use of ﬂow cytometry [116] to evaluate

cell markers could be considered to be comparable to the use of histologic markers, and

involves the extraction of biologic samples to evaluate cell distribution.

4. Discussion

Determining the fate of MSCs after administration is a major issue in the development

of cell therapies. On one hand, as a part of their physiological functions, MSCs are able

to produce several soluble substances and to modulate the immune response through

different pathways; the production and induction of interleukin production and the release

of microvesicles [123–125]. Cell interactions lead to the secretion of soluble factors and

cell-to-cell contact which induces changes in the immunobiology of immune cells, such as

changing the interleukin production, inducing anergy or triggering apoptosis. On the other

hand, MSCs have been found to be able to differentiate into different cellular subtypes,

which could play a role in regenerative medicine. Whether MSCs act by modulating the

immune system or differentiating into tissular cells, understanding how and where cells

are distributed after being administered by each route of administration is critical.

Intravenous injection of MSCs remains the most widely used form of administration.

The widespread use of this route of administration for drugs which are used in clinical

practice, and the ease of administering cells by this route, are probably the reasons why.

As previously seen, IV administration might be beneﬁcial when cell trapping in the lungs,

liver or spleen is pursued, or when MSCs are capable of acting at a distance. However,

intraarterial delivery might be of choice when wide systemic distribution into different

tissues and organs is required. Moreover, if a targeted deposition of cells into a single

organ is needed, intraarterial selective delivery could be the solution. In contrast to

intravascular administration, intramuscular injection seems to lack signiﬁcant systemic

distribution of cells and might be preferred when the cells do not necessarily need to reach

the target tissues.

Regarding intralesional administration of MSCs, there are several distribution patterns

depending on the organ or tissue injected. Intraarticularly injected MSCs seem to remain

in the joints, which could be of beneﬁt when treating articular diseases. Administration

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into skin, lymph nodes, trachea, lungs and urinary tissues does not produce systemic

distribution either, and might be useful when targeted delivery is required. In contrast,

intrahepatic, intrasplenic, intracardiac and intrapericardial infusion led to a distribution of

MSCs following the natural direction of the bloodstream. Moreover, the injection of cells

into virtual anatomical cavities containing corporal ﬂuids seems to produce a distribution

of MSCs within the same anatomic areas: intraperitoneal, intra-cerebro-ventricular and

intrathecal routes of administration make MSCs reach the organs and tissues in contact

with the correspondent ﬂuid. Finally, intracerebrally administered MSCs are able to move

within the brain if induced to do so by appropriate stimuli.

5. Limitations and Future Studies

Although the fate of MSCs after each route of administration seems to be fairly well

understood, the speciﬁc mechanisms which lead to these distribution patterns are still a

matter of discussion. Moreover, as has been previously reviewed [30,46], these mechanisms

might be modulated by speciﬁc factors such as the surface molecules expressed by MSCs

or the age of the donors and recipients of cells. Although this is still unknown, other

factors, such as the speciﬁc subtype of MSC or the donor and recipient model, could also be

important. Moreover, there is great variability among different studies with respect to the

exact forms of administration (e.g., the exact anatomical site injected or the concentration

or volume of cells administered). The design of standardized protocols for mesenchymal

cell administration could lead to less variability of results, making them more comparable.

6. Conclusions

Biodistribution of MSCs in animal models and humans appears to be comparable. In

response to the research questions, some facts are worth noting:

(a)

Intravenous administration leads to an initial accumulation of cells in the lung with

later redistribution to the liver, spleen and kidneys;

(b)

Intraarterial injection bypasses the pulmonary ﬁlter, so MSCs distribute more widely

into the rest of the organs of the body;

(c)

In both of the two previous routes of administration, selective perfusion of selected

blood vessels is useful for targeting speciﬁc organs;

(d)

MSCs are not distributed systemically in signiﬁcant quantities after intramuscular,

intraarticular, intradermal, intranodal, intratracheal, intrapulmonary and intraurinary

tissue administration;

(e)

The injection into speciﬁc organs, such as the liver, spleen, pericardium or heart leads

to a distribution of MSCs following the direction of the natural bloodstream;

(f)

The injection into anatomical cavities containing body ﬂuids (cerebral ventricles,

subarachnoid space and peritoneum) leads to a distribution of MSCs in tissues which

are in contact with the ﬂuid;

(g)

MSCs injected intracerebrally seem to be able to migrate within the central ner-

vous system.

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Table 2. Biodistribution after IV administration of MSCs in animal models.

Article

Model

Disease (Number of

Animals)

Route of

Administration

(Source of Cells)

Cell-Marking Technique

Detection Time and Outcome

Comments

Krueger et al.

[126] (2018)

Adult

baboons [26]

Lethal total body

irradiation

(3 animals)

Intravenous

(autogenic and

allogenic MSCs)

Genetic transduction with green

ﬂuorescent protein retroviral

construct, which was later

evaluated by PCR.

Necropsies were performed between 9 and

21 months following MSC infusion.

Several tissues were found to have MSCs:

Gastrointestinal, kidney, skin, lung,

thymus, and liver.

Gastrointestinal tissues had

the highest MSCs

concentration.

MSCs distribute to a wide

variety of tissues following

systemic administration.

Mongrel dogs

[24]

Miocardial infarction

(7 animals)

Intravenous

(allogenic MSCs)

111In oxine–labeled MSCs

colabeled with

ferumoxides–poly-l-lysine.

Single-photon emission CT

(SPECT) and x-ray CT

(SPECT/CT) and MRI studies

were used to evaluate the

distribution.

Imaging was performed immediately after

injection and at multiple time points

between 1 and 7 days after infusion.

Early imaging showed a high distribution

to lungs, which later decreased drastically.

After day 1, MSCs distributed from lungs

to different organs (kidney, bone marrow,

liver, spleen) and also to the infarcted area.

A high and early

distribution to lungs is

showed, with a progressive

decrease of MSCs and a later

redistribution to a wide

variety of tissues.

Mice [25]

Miocardial infarction

(number unknown)

Intravenous

(xenogenic

MSCs—human

MSCs)

Human MSCs were infused,

Quantitative assays for human

DNA and mRNA were used to

evaluate the distribution,

Tests were done at 15 min, and up to 100 h

post infusion.

Early distribution to the lungs was detected

(15 min).

Later distribution to other organs was

insigniﬁcant: less than 1% of cells was

detected in any other organ after 48 h.

Authors conclude that

effects of intravenous MSCs

might be due to soluble

mediators rather than

engraftment of MSCs in

target tissues.

Mello et al.

[39] (2020)

Rats

Intracerebral

hemorrhage

Intravenous

(xenogenic

MSCs—human

MSCs)

99mTc was used to label MSCs.

Scintigraphy and radioactivity

measurements (cerebral

hemispheres, heart, lungs, liver,

kidneys, intestines, and spleen)

were performed to assess

biodistribution.

Scintigraphy was performed 2 h after cell

injection and ex vivo radioactivity was

evaluated 24 h after cell transplantation.

MSCs were mainly distributed to the lungs,

kidneys, spleen and liver. Brain captation

was low but it was relatively higher in the

damaged hemisphere.

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Table 2. Cont.

Article

Model

Disease (Number of

Animals)

Route of

Administration

(Source of Cells)

Cell-Marking Technique

Detection Time and Outcome

Comments

Patrick et al.

[127] (2020)

Mice

Lung cancer

Intravenous

(xenogenic

MSCs—human

MSCs)

89Zr-oxine and luciferase were

used to label MSCs. PET-CT,

bioluminescence and ex vivo

radioactivity measures were

used to assess biodistribution.

PET-CT at 1 h and 1, 2, and 7 days

post-injection. At 7 days, radioactivity was

measured from ex vivo organs.

The majority of signal (60%) was found in

the lung at 1 h before decreasing, while

liver signal increased. From 1 to 7 days

post-injection, the proportion of the ⁸⁹Zr

signal in the lung fell further from 24.6%.

Wuttisarnwattana

et al. [128]

(2020)

Mice

Bone marrow

transplanted animals

Intravenous

(xenogenic

MSCs—human

MSCs)

Red quantum dots were used to

label MSCs. Ex vivo

cryo-imaging was performed to

assess biodistribution in

different tissues (lung, liver,

spleen, kidneys, bone marrow).

Animal sacriﬁce was performed at different

time points following stem cell infusion (24,

48, 72 h).

Initially, MSCs were found as clusters in the

lung and eventually dissociated to single

cells and redistributed to other organs

within 72 h, mainly to the spleen and liver.

De White

et al. [32]

(2018)

Mice

Healthy animals

(number unknown)

Intravenous

(xenogenic

MSCs—human

MSCs)

Qtracker 605 beads and

Hoechst33342, which labelled

alive and dead cells, respectively.

Anatomical and molecular

ﬂuorescence videos were

generated with CryoViz

Technology.

Blood tests were performed to

analyze phagocytosis.

Necropsies were performed at 5 min, 24 h

and 72 h post-infusion.

Early accumulation of MSCs in the lungs

(5 min) was demonstrated. MSCs were

phagocytized in the lungs and redistributed

to liver within the monocytes at 24 and 72 h.

Monocytes change their immunophenotype

after phagocyting MSCs, and induce Treg

cells.

Authors conclude that the

action of MSCs in many

organs may be due to the

phagocytosis of MSCs by

monocytes and the later

change in their phenotype,

which leads to the induction

of Treg cells.

Ehrhart et al.

[35] (2016)

Mice and rats

Alzheimer’s disease

model

Intravenous

(xenogenic

MSCs—human

MSCs)

Human MSCs were used.

Tisular PCR analyses (blood,

bone marrow, brain, spinal cord,

spleen, kidney, liver, heart, lung,

gonad) were used to assess

biodistribution.

Harvesting of tissues was performed at

24 h, 7 days, and 30 days after injection.

MSCs were broadly detected both in the

brain and several peripheral organs,

including the liver, kidney, and bone

marrow, of both species, starting within

7 days and continuing up to 30 days

post-transplantation.

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Table 2. Cont.

Article

Model

Disease (Number of

Animals)

Route of

Administration

(Source of Cells)

Cell-Marking Technique

Detection Time and Outcome

Comments

Tang et al.

[129] (2016)

Rats

Cirrhosis rats

(splenectomized)

Intravenous

(allogenic MSCs)

Qtracker705

nanoparticle-labelled MSCs

were infused.

Fluorescence imaging was

performed to assess

biodistribution.

Images were taken at 2 h and 5 days after

cell infusion.

Splenectomy improved the homing of

MSCs in the liver when compared to

non-splenectomy group.

Cao et al.

[130] (2016)

Rats

Healthy animals

Intravenous

(allogenic MSCs)

Luciferase and green ﬂuorescent

protein were used to label MSCs.

Bioluminescence imaging, ex

vivo organ imaging,

immunohisto-chemistry (IHC),

and RT-PCR were used to assess

biodistribution.

Images were taken up to 1 month. After

that, histological analysis was performed.

MSCs were detected initially in the lungs

with subsequent distribution to liver,

kidneys and other abdominal organs. The

dorsal skin was also detected to have MSCs.

The signals disappeared at day 14.

Zhou et al.

[131] (2015)

Rats

Hepatic ﬁbrosis

Selective intravenous

(superior mesenteric

vein)

(allogenic MSCs)

MSCs were double-labeled with

superparamagnetic iron oxide

and green ﬂuorescent protein.

MRI, histology and qPCR tests

were used to assess

biodistribution.

MR imaging of the liver was carried out

before and 1, 3, 7 and 12 days after injection.

Liver, lung, kidney, muscle and heart

tissues were harvested at 1, 7, 15 and

42 days after cell injection.

Dual-labeled MSCs were retained in the

ﬁbrotic liver of rats. SPIO particles and

EGFP-labeled BMSCs showed a different

tissue distribution pattern in rats with liver

ﬁbrosis at 42 days after transplantation.

SPIO-based MR imaging

may not be suitable for

long-term tracking of

transplanted BMSCs in vivo.

Kim et al. [36]

(2015)

Mice

(athymic)

Brain tumor

Intravenous and

intracerebral

(xenogenic

MSCs—human

MSCs)

MSCs were labeled with

near-infrared ﬂuorescent dye.

Bioluminescence and

ﬂuorescence imaging, qPCR and

histologic examinations were

performed.

Imaging techniques were performed at 1

and 4 h, 1, 7, 14 and 21 days.

MSCs resided predominantly in the lung

up to day 1 and the signal intensity

decreased over time. Many cells moved

from the lung toward other organs (liver

and spleen) after day 1, and the signal

remained stable in these regions for 14 days.

From day 1 to day 14, MSCs were clearly

detectable in the tumor area.

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Table 2. Cont.

Article

Model

Disease (Number of

Animals)

Route of

Administration

(Source of Cells)

Cell-Marking Technique

Detection Time and Outcome

Comments

Kim et al. [38]

(2015)

Beagle dogs

Spinal cord injury

Intravenous

(allogenic MSCs)

MSCs were labeled with green

ﬂuorescent protein.

Ex vivo bioluminescence was

used to assess biodistribution.

Ex vivo examination was performed 7 days

after injection.

The green ﬂuorescent protein-expressing

AD-MSCs were clearly detected in the lung,

spleen, and injured spinal cord; however,

these cells were not detected in the liver

and un-injured spinal cord.

Li et al. [45]

(2015)

Mice

Acute liver injury

Selective

intravenous: Inferior

vena cava (IVC),

superior mesenteric

vein (SMV) and

intrahepatic (IH)

injection.

(allogenic MSCs)

MSCs were labeled with

luciferase. Bioluminiscece

images were used to assess

biodistribution.

Images were taken at 3 h, and at 1, 3, 7, 10,

14 and 21 days.

After IVC infusion, MSCs were quickly

trapped inside the lungs, and no detectable

homing to the liver was observed. By IH

injection, lung entrapment was bypassed,

but MSCs-R distribution was only localized

in the injection region of the liver. After

SMV infusion, MSCs-R were dispersedly

distributed and stayed as long as 7-day

post-transplantation in the liver.

SMV is the optimal MSCs

delivery route for liver

disease.

Zhang et al.

[33] (2015)

Rabbit

Acute ischemic

kidney injury

Intravenous

(allogenic MSCs)

MSCs were labeled with SPION

particles. MRI images and

histological analysis were used

to assess biodistribution

Images and histological analysis were

taken at 1, 3, 5 and 8 days.

MSCs were detected up to 8 days, with a

maximum amount of cells at day 3.

No systemic distribution was assessed.

Schmuck et al.

[43] (2016)

Sprague-

Dawley

rats

Acute lung injury

(12 animals)

Intravenous

(xenogenic

MSCs—human

MSCs)

MSCs were labeled with

QTracker65. 3D cryo-imaging of

lungs, liver, spleen, heart,

kidney, testis, and intestine was

performed to assess

biodistribution.

Tissue samples were collected and

analyzed at 60, 120 and 240 min and 2, 4

and 8 days after infusion.

Distribution up to 240 min was detected

mostly in liver, and also in lungs and

spleen.

The number of cells detected at 2, 4, and

8 days was less than 0.06% of the total cells

infused on day 0 and were mainly

distributed also in lungs, liver and spleen

but relatively higher captation was seen in

the rest of the tissues studied.

Authors conclude that

studies using

bioluminescence to track

cells underestimate cell

retention in the liver because

of its high tissue absorption

coefﬁcient

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Table 2. Cont.

Article

Model

Disease (Number of

Animals)

Route of

Administration

(Source of Cells)

Cell-Marking Technique

Detection Time and Outcome

Comments

Li et al. [27]

(2018)

Rats

Silicosis

(54 animals)

Intravenous

(allogenic MSCs)

MSCs were labelled with

1,1^′-dioctadecyltetramethyl

indotricarbocyanine iodide.

Fluorescence imaging was

performed to assess

biodistribution.

Images were taken 1 h, 6 h, 24 h, 3 days,

15 days, and 30 days after injection both

in vivo and ex vivo.

MSCs distributed mostly in liver and lungs,

with a peak at 6 h, and a dramatic decrease

by day 3. At day 30, no MSCs were

detected.

Distribution in lungs was

signiﬁcantly higher in rats

with damaged lungs

compared to healthy rats.

Park et al.

[34] (2018)

Mice

Alzeimer’s disease

(53 animals)

Intravenous

(allogenic MSCs)

MSCs were ¹¹¹In-tropolone

labeled. Imaging with SPECT

(in vivo) and gamma-counter

(ex vivo) was performed to

assess biodistribution.

Imaging and gamma-counter studies were

performed at 24 h and 48 h post infusion.

In Alzheimer’s model, brain uptake of

MSCs was signiﬁcantly higher than in

healthy animals. In both groups, MSCs

distributed mainly to lungs, liver and

spleen.

Distribution to brain seem to

be higher in Alzheimer’s

models.

Leibacher

et al. [28]

(2017)

Mice

Healthy animals

(number unknown)

Intravenous

(xenogenic

MSCs—Human

MSCs)

Human MSCs were injected and

PCR techniques were used to

assess biodistribution by

searching for SRY sequences.

Ex vivo assessment was performed at

5 min, 30 min, 2 h, 6 h, and 24 h.

The majority of injected MSCs were

detected by qPCR in the lungs 5 min after

transplantation, whereas <0.1% were

detected in other tissues over 24 h

After intravenous injection,

most cells distribute to

lungs.

Yun et al. [31]

(2016)

Rats

Acute liver injury

Intravenous

(xenogenic

MSCs—Human

MSCs)

Human MSCs were injected and

PCR techniques were used to

assess biodistribution.

Mice were euthanized at 1, 3, 12, or 24 h

and at 1, 4, or 13 weeks post injection.

MSCs were detected soon in the lungs and

disappeared before 1 week post injection.

Then, MSCs were found mainly in the liver.

No MSCs were found in other tissues

(testis, ovary, spleen, pancreas, kidney,

adrenal gland, thymus, and brain).

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Table 2. Cont.

Article

Model

Disease (Number of

Animals)

Route of

Administration

(Source of Cells)

Cell-Marking Technique

Detection Time and Outcome

Comments

Abramowski

et al. [42]

(2016)

Mice

Experimental

autoimmune

encephalomyelitis

model

(number unknown)

Intravenous

(allogenic MSCs)

MSCs were injected and a

variety of techniques, including

magnetic resonance imaging,

immunohistochemistry,

ﬂuorescence in-situ

hybridization, and quantitative

polymerase chain were

performed to assess

biodistribution.

Assessment was focalized in the brain area.

No evidence for immediate migration of

infused MSC into the central nervous

system of treated mice was found.

Kim et al. [30]

(2016)

Rats

Healthy rats

Intravenous

(allogenic MSCs)

MSCs were surface-modiﬁed

with HA—wheat germ

agglutinin (WGA) conjugate for

targeted systemic delivery of

MSCs to the liver and labeled

with ﬂuorescent dyes.

Histologic examinations were

performed.

Assessment was performed at 4 h post

injection. Lungs and livers were collected.

HA-WGA-MSCs had a greater distribution

to the liver when compared to control

MSCs, which were mainly trapped in the

lungs.

HA-WGA conjugate has

great potential to deliver

MSCs to the liver efﬁciently

within a short time and to

reduce the entrapment of

MSCs in the lung.

Lu et al. [40]

(2016)

Mice

Acute distress

respiratory

syndrome model

Intravenous

(allogenic MSCs)

Fluorescein isothiocyanate–

dextran was used to label MSCs.

Histological analyses and qPCR

were used to assess

biodistribution.

Assessment was performed immediately

after cell injection, 2, 24, and 48 h later.

Lung, heart, spleen, kidney, brain, and liver

were collected.

MSCs accumulated mainly in the lungs of

control and diseased mice, with minor

amounts distributed to other organs up to

2 h. Diseased animals showed less early

distribution to lungs and higher

distribution to the rest of the organs when

compared to healthy animals.

Acute distress respiratory

syndrome might lessen the

pulmonary capillary

occlusion by MSCs

immediately following cell

delivery while facilitating

pulmonary retention of the

cells.

J. Clin. Med. 2021, 10, 2925

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Table 2. Cont.

Article

Model

Disease (Number of

Animals)

Route of

Administration

(Source of Cells)

Cell-Marking Technique

Detection Time and Outcome

Comments

Fabian et al.

[46] (2017)

Young and

old mice

Alzheimer disease

(unknown number)

Intravenous

(syngenic MSCs)

Histologic and genetic tests

(PCR) were performed to

evaluate MSCs distribution.

Genetic tests and histology were assessed

after 28 days.

Transplantation of MSCs obtained from old

mice showed biodistribution only in the

blood and spleen in both young and old

mice.

MSCs obtained from young mice showed a

wide distribution in young receptors (lung,

axillary lymph nodes, blood, kidney, bone

marrow, spleen, liver, heart, and brain

cortex). In contrast, these cells showed

distribution only in the brain cortex in old

mice.

Authors conclude that aging

of both the recipient and the

donor MSCs used attenuates

transplantation efﬁciency.

Ohta et al.

[37] (2017)

Rats

Spinal cord injury

Intravenous

(allogenic MSCs)

MSCs were labeled with

3H-thymidine. Histologic and

radioactivity examination of the

spinal cord segment containing

the damaged region, blood and

target organs were harvested.

After 3, 24 and 48 h, organs were collected

and radioactivity measured.

The highest radioactivity was detected in

the lungs 3 h after infusion, while

radioactivity in the injured spinal cord was

much lower. However, brain radioactivity

was lower than damaged spinal cord.

MSCs distribute to the

injured spinal crod.

Liu et al. [29]

(2018)

Mice

Acute lung injury

Intravenous

(xenogenic

MSCs—Human

MSCs)

MSCs were labeled with

ﬂuorophore Cy7.

Histology was performed to

assess biodistribution.

Ex vivo assessment of lungs, heart, spleen,

kidneys and liver was performed at 30 min,

1 day, 3 days and 7 days following injection.

MSCs distributed to the lungs up to day 1;

and to the liver up to day 3, with

progressive subsequent decrease. No

signiﬁcant distribution was observed to

heart, spleen and kidneys

J. Clin. Med. 2021, 10, 2925

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Table 2. Cont.

Article

Model

Disease (Number of

Animals)

Route of

Administration

(Source of Cells)

Cell-Marking Technique

Detection Time and Outcome

Comments

Qin et al. [41]

(2018)

Rabbits

Liver tumors

Intravenous

(allogenic MSCs)

MSCs were colabeled with

superparamagnetic iron oxide

(SPIO) particles and

4^′,6-diamidino-2-phenylindole

(DAPI).

MRI and histologic examination

were performed.

MRI was performed at days 0, 3, 7 and 14

after cells transplantation. Histological

analyses were performed immediately after

the MRI examination.

MSCs were detected in the liver tumors,

rather than the non-tumor liver tissue and

other organs. At day 3, MSCs were mainly

in the central part of the tumor, showing a

posterior distribution in the periphery.

MSCs distribute mainly to

the damaged liver when

injected intravenously.

Leibacher

and

Henschler

[132] (2016)

Wistar rats

[133]

Transient cerebral

ischemia

(25 animals)

Intravenous and

intraarterial

(allogenic MSCs)

Feridex (Berlex Imaging) mixed

with the transfection agent

poly-l-lysine.

Later evaluation with MRI and

necropsies.

Imaging was performed before and after

the infusion (2 to 24 h after).

After intraarterial infusion, MSCs were

detected in the brain of the rats.

After intravenous infusion, no MSCs were

detected in the brain.

Authors conclude that MSCs

may engraft in peripheral

tissues after intraarterial

infusion. Intravenous

infusion might not be quite

effective to deliver MSCs to

peripheral tissues.

Mice [47]

Healthy animals and

acute kidney injury

(AKI) model

(Unknown number)

Intravenous and

intraarterial.

(allogenic MSCs)

Transfection with

luciferase-neomycin

phosphotransferase construct.

Later evaluation with Xenogen

IVIS 100 imaging system.

Imaging was performed immediately after

infusion, at 24 h, 72 h and 7 days.

Intravenous infusion led to a majority of

cells distributing to lungs.

Intraarterial infusion lacked pulmonary

retention and caused distribution to

kidneys, especially in AKI mice. MSCs

gradually disappeared after 24 h.

Intraarterial infusion might

be adequate when treating

kidney conditions.

Schubert et al.

[44] (2018)

Mice

Acute kidney injury

model

(Unknown number)

Intravenous.

(autogenic MSCs)

MSCs from luciferase transgenic

mice.

Evaluation was performed with

bioluminescence imaging and

RT-PCR.

Imaging was performed on days 1, 3 and 6.

RT-PCR was performed in kidney, lung,

liver tissue and blood on day 6.

Bioluminescence showed a high

distribution of MSCs to lungs on day 1,

which disappeared on days 3 and 6.

RT-PCR on day 6 showed variables

amounts of MSCs-mRNA in blood, liver

and kidneys

RT-PCR seems to be a more

sensitive technique to

demonstrate the late

presence of MSCs in

different tissues when

compared to

bioluminescence.

J. Clin. Med. 2021, 10, 2925

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Table 2. Cont.

Article

Model

Disease (Number of

Animals)

Route of

Administration

(Source of Cells)

Cell-Marking Technique

Detection Time and Outcome

Comments

Nakada and

Kuroki [62]

Mice

Healthy animals

(Unknown number)

Intravenous and

intramuscular

(allogenic MSCs)

MSCs were labelled with

chromium.

Laser ablation inductively

coupled plasma imaging mass

spectrometry (LAICP-IMS) was

used to assess biodistribution,

Detection time is not recorded.

After intramuscular injection, MSCs remain

in the muscular tissue.

After intravenous injection, MSCs are

detected in the lungs.

Authors conclude that

chromium labelling could be

a promising technique.

Mäkelä et al.

[22] (2015)

Pigs

Healthy animals (12

animals)

Intravenous and

intraarterial

(autogenic and

allogenic MSCs)

99mTc- hydroxymethyl-

propylene-amine-oxime.

Evaluation was performed with

SPECT/TC. Biopsies were also

performed.

Imaging was performed 8 h later.

Intravenous infusion led to a high

distribution of MSCs into the lungs.

Intraarterial infusion decreased the

deposition in the lungs and increased the

uptake in other organs, specially the liver

and kidneys.

Intraarterial infusion might

improve the distribution to

peripheral tissues and may

avoid pulmonary retention.

Wang et al.

[134] (2015)

Mice

Bone marrow

transplanted animals

Intravenous and

intraarterial

(xenogenic

MSCs—Human

MSCs)

99mTc- hydroxymethyl-

propylene-amine-oxime and

luciferase.

Bioluminescence, scintigraphy

and histologic examination were

used to assess biodistribution.

Bioluminescence was performed at 30 min,

24 h, 48 h, 96 h and once a week for up to

two month. Scintigraphic imaging and

X-ray imaging were performed at 5 h, 10 h

and 1 d after injection. After 2 months,

animals were sacriﬁced and ex vivo

histology was performed.

After intraarterial injection persistent

whole–body MSC distribution in

allo-trasplant recipients was shown, while

MSCs were rapidly cleared in the syngeneic

animals within one week. In contrast,

intravenous injected MSCs were mainly

seen in the lungs with fewer cells traveling

to other organs.

J. Clin. Med. 2021, 10, 2925

24 of 51

Table 2. Cont.

Article

Model

Disease (Number of

Animals)

Route of

Administration

(Source of Cells)

Cell-Marking Technique

Detection Time and Outcome

Comments

Silachev et al.

[49] (2016)

Rats

Traumatic brain

injury model

Intravenous and

intraarterial

(allogenic MSCs)

9mTc and iron microparticles

labelled MSCs. Evaluation was

performed with SPECT/TC,

MRI and histology.

Evaluation was performed at 1 h and 16 h

after trasplantateion.

After intravenous injection, MSCs

distributed to lung, kidney, and partially in

the liver and bladder, with progressive

decrease to 16 h. After intraarterial

injection, MSCs distributed signiﬁcantly to

damaged hemisphere.

Intraarterial injection

improves the distribution to

the damaged cerebral area.

Cao et al. [50]

(2018)

Rats

Orthotopic glioma

model

Intravenous,

intraarterial and

intratumoral

(allogenic MSCs)

MSCs were transduced to

express ferritin heavy chain and

green ﬂuorescent protein.

MRI and histology evaluations

were performed.

MRI was performed at days 0, 1, 3, 5, 7 and

9 after cell injection. Histological analysis

was performed at days 8, 12 and 18.

Intravenous injection did not lead to

accumulation of MSCs in the tumor.

However, intralesional and intraarterial

injections showed a rapid accumulation of

MSCs in the core of the tumor with a

gradual decrease of the cells in the zone.

Intravenous injections does

not lead to MSCs migration

to central nervous system

tumors, whereas

intraarterial and

intralesional injections do.

Taylor et al.

[55] (2020)

Mice

Renal injury model

Intravenous and

intracardiac

(allogenic MSCs)

MSCs were labelled with

luciferase and SPIO. MRI and

bioluminescence were used to

assess biodistribution.

Images were taken up to 2 days after

injection.

Following intravenous administration, no

MSCs were detected in the kidneys,

irrespective of whether the mice had been

subjected to renal injury. After intracardiac

injection, MSCs transiently populated the

kidneys, but no preferential homing or

persistence was observed in injured renal

tissue.

J. Clin. Med. 2021, 10, 2925

25 of 51

Table 2. Cont.

Article

Model

Disease (Number of

Animals)

Route of

Administration

(Source of Cells)

Cell-Marking Technique

Detection Time and Outcome

Comments

Scarfe et al.

[48] (2018)

Mice

Healthy animals

(unknown number)

Intravenous and

intracardiac (left

ventricle)

(allogenic MSCs and

xenogenic

MSCs—human

MSCs)

MSCs were labelled with

luciferase (Luc) or a bicistronic

construct of Luc and ZsGreen

for bioluminescence imaging.

For MR tracking, cells were

labelled with diethylaminoethyl-

dextran-coated

SPIONs.

In vivo biodistribution of cells was

monitored by BLI immediately after cell

administration and at multiple time points

up to 30 day. Ex vivo MRI at baseline and

up to 2 days post administration.

Intravenous MSCs distributed mainly to

the lungs.

Intracardiac MSCs distributed to the brain,

heart, lungs, kidney, spleen and liver, with

also a majority of cells distributing to the

lungs.

Intracardiac injection led to

a wide distribution of MSCs

to peripheral organs.

Table 3. Biodistribution after IA administration of MSCs in animal models.

Article

Model

Disease

(Number of

Animals)

Route of

Administration

(Source of Cells)

Cell-Marking Technique

Detection Time and Outcome

Comments

Khabbal et al.

[51] (2015)

Rats

Ischemic stroke

model

Intraarterial (external

carotid)

(allogenic MSCs and

xenogenic

MSCs—Human

MSCs)

MSCs were labeled with ^99mTc.

Whole body SPECT images and ex

vivo radioactivity measures were

used to assess biodistribution.

SPECT images were acquired 20 min, 3 h, and 6 h

postinjection, after which rats were sacriﬁced for ex

vivo examinations.

The majority of the cells were located in the brain

and especially in the ipsilateral hemisphere

immediately after cell infusion. This was followed

by fast disappearance. At the same time, the

radioactivity signal increased in the spleen, kidney,

and liver.

Human MSCs had faster

clearance from the brain

than rats MSCs.

Fukuda et al.

[56] (2015)

Rats

Ischemic stroke

model

Intraarterial

(Common carotid

artery)

(xenogenic

MSCs—human

MSCs)

Human MSCs were used and

labeled with PKH26.

Bioluminescence and anti-human

vimentin antibodies were used to

assess biodistribution of MSCs in ex

vivo histological analysis.

Examinations were performed 24 h post infusion.

MSCs were widely distributed throughout the

cortex and striatum of the ipsilateral hemisphere at

24 h after transplantation of MSCs.

J. Clin. Med. 2021, 10, 2925

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Table 3. Cont.

Article

Model

Disease

(Number of

Animals)

Route of

Administration

(Source of Cells)

Cell-Marking Technique

Detection Time and Outcome

Comments

Cerri et al.

[52] (2015)

Wistar

rats

Parkinson’s

disease

(unknown

number)

Intraarterial. (One

group also received

mannitol to

transiently

permeabilize the

blood-brain barrier).

(allogenic MSCs)

MSCs were double-labelled:

CellVue NIR815 Kit for Membrane

Labeling (Polyscience, Warrington,

PA, http://www.polysciences.com)

(accessed on 25 June 2021)

and lipophilic red ﬂuorescence dye

PKH26. Later histological

examinations assessed the

distribution of MSCs within the

brain.

Necropsies were performed 7 and 28 days after

infusion of MSCs.

Rats not treated with mannitol showed a very low

number of MSCs in the brain at 7 and 28 days

post-infusion. Rats treated with mannitol showed

a signiﬁcantly higher number of MSCs within the

brain. At day 7, most of MSCs were in the blood

vessels, whereas at day 28, most of MSCs were in

the parenchyma.

Most of MSCs distributed in the same lateral

hemisphere where the infusion took place.

A strong MSCs signal in the lungs and spleen up to

28 days after infusion was detected.

Authors conclude that the

use of a permeabilizing

agent is essential to allow

passage of MSCs across the

blood-brain barrier.

A signiﬁcant number of

infused cells accumulated in

the peripheral organs (liver,

lungs).

Jin et al. [135]

(2016)

Beagle

dogs

Osteonecrosis of

the femoral head

Intraarterial

(autogenic MSCs)

MSCs were labeled with

5-bromo-2-deoxyuridin. Histologic

examinations (right femoral head,

heart, lung, liver, spleen, kidney,

gallbladder, small bowel, pancreas,

prostate, and testicle) were

performed to assess biodistribution.

Histologic examinations were performed 8 weeks

after cell infusion.

Organs had uneven distribution of MSCs: Heart,

liver, gallbladder, kidney and stomach had the

major quantity of MSCs.

Arnberg et al.

[58] (2016)

Rabbit

Healthy rabbits

Intraarterial infusion

(superior mesenteric

artery) and

intravenous

(xenogenic

MSCs—Human

MSCs)

MSCs were labeled with

11In-oxinate.

SPECT-TC images were used to

assess biodistribution.

SPECT-TC was performed at 6 h and at 1, 2, and

5 days post infusion.

Intravenous administration resulted in early and

long distribution of MSCs to the lungs. In contrast,

selective intraarterial injections resulted in MSCs

distribution in the intestine and in the liver.

Selective intraarterial

delivery could improve the

results in treating some

localized diseases.

Espinosa et al.

[59] (2016)

Horses

Healthy horses

Intraarterial selective

infusion (median

artery)

(allogenic MSCs)

MSCs were labeled with

99mTc-HMPAO. Scintigraphic

images were taken to assess

biodistribution.

Images were taken at the time of injection and at 1,

6, and 24 h postinjection.

Homogeneous distribution of radiolabeled MSC

was observed through the entire distal limb,

including within the hoof. Systemic

biodistribution was not assessed.

J. Clin. Med. 2021, 10, 2925

27 of 51

Table 3. Cont.

Article

Model

Disease

(Number of

Animals)

Route of

Administration

(Source of Cells)

Cell-Marking Technique

Detection Time and Outcome

Comments

Sierra-

Parraga et al.

[57] (2019)

Pigs

Renal ischemia-

reperfusion

injury.

(unknown

number).

Intraarterial infusion

(renal artery)

(allogenic MSCs)

MSCs were labelled with

ﬂuorescent compunds. Flow

cytometry and genetic tests (PCR)

were done in blood and tissues.

Samples were collected 30 min and 8 h after

infusion.

After infusion, a minor number of MSCs left the

kidney through the renal vein, and no MSCs were

identiﬁed in arterial blood. A low percentage of

the infused MSCs were present in the kidney

14 days after administration.

Most of MSCs were trapped in the renal cortex.

Renal intra-arterial MSC

infusion seem to limit

off-target engraftment,

leading to efﬁcient MSC

delivery to the kidney.

Barthélémy

et al. [60]

(2020)

Golden

Re-

triever

Dogs

Duchenne

muscular

dystrophy model

Intraarterial (femoral

artery)

(not stated)

MSCs were labeled with ¹¹¹In-oxine.

Scintigraphy was performed to

assess biodistribution.

Scintigraphic images were taken immediately after

injection and at 1, 2, 24, 48 h and 1 week.

Immediately after injection, MSCs were trapped in

the capillary network of the limb and in the lungs.

Subsequently, MSCs were also mainly in the

injected limb, with a decrease in the lung captation

and a relative increase in the liver captation.

Table 4. Biodistribution after intramuscular and intraarticular administration of MSCs in animal models.

Article

Model

Disease

(Number of

Animals)

Route of

Administration

(Source of Cells)

Cell-Marking Technique

Detection Time and Outcome

Comments

Hamidian

Jahromi et al.

[63] (2017)

Mice

Carrageenan-

induced plantar

inﬂammation

Intramuscular

(contralateral to

plantar

inﬂammation)

(xenogenic

MSCs—Human

MSCs)

MSCs were labelled with Gaussia

Luciferase.

Bioluminescence imaging, qPCR

and histology techniques were used

to assess biodistribution.

Bioluminescence was performed at 24 h, 48 h and

up to 33 days.

No MSCs were found to distribute to other organs.

MSCs were detectable in the muscle up to 33 days

after injection.

MSCs were able to reduce

the contralateral

inﬂammation and to lower

the TNF-alfa serum levels

without distributing

systemically.

J. Clin. Med. 2021, 10, 2925

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Table 4. Cont.

Article

Model

Disease

(Number of

Animals)

Route of

Administration

(Source of Cells)

Cell-Marking Technique

Detection Time and Outcome

Comments

Creane et al.

[61] (2017)

Mice

Healthy mice

(10 animals)

Intramuscular

(xenogenic

MSCs—Human

MSCs)

Human MSCs were injected and

quantitative PCR for Alu sequences

was performed in different tissue

samples.

Ex vivo analysis was performed 3 months after

injection.

No MSCs were detected in any organ, including

heart, lung, brain, liver, kidney and spleen. MSCs

were detected in the thigh and calf samples, where

MSCs were injected.

Intramuscular MSCs do not

seem to remain viable

and/or distribute 3 months

after injection.

Hamidian

Jahromi et al.

[65] (2019)

Rats

and

mice

(Re-

view)

Different diseases

Intramuscular

(different sources of

MSCs)

Different techniques.

MSCs do not seem to distribute after intramuscular

injection. MSCs seem to remain or spread locally,

without systemic biodistribution.

Intramuscular MSCs do not

seem to distribute

systemically.

Cai et al. [64]

(2017)

Rats

Healthy rats

Intramuscular

(allogenic MSCs)

Melanin-based gadolinium3+

(Gd3+)-chelate nanoparticles were

used to label MSCs.

MRI was used to assess

biodistribution.

MRI was performed on days 1, 4, 7, 14, 21, and 28.

MSCs were found in the muscle up to 28 days after

injection. No systemic biodistribution was

observed.

Intramuscular MSCs do not

seem to distribute

systemically

Markides

et al. [70]

(2019)

Sheep

Osteochondral

injury

Intraarticular

(autogenic MSCs)

MSCs were labelled with Nanomag,

and using a cell-penetrating

technique,

glycosaminoglycan-binding

enhanced transduction (GET).

Evaluation was performed with ex

vivo MRI and histologic tests.

Ex vivo MRI and histology was performed 7 days

after injection.

MSCs were detected in the synovium, and not in

the osteochondral defect.

MSCs are capable to home in

the synovium, whereas they

do not seem to be able to

enter the joint to reach the

osteochondral defect.

Yang et al.

[74] (2019)

Mice

Supraspinatus

tendon tear

Intraarticular

(allogenic MSCs)

MSCs were labeled with quantum

dots with near-infrared properties.

Near-infrared ﬂuorescence imaging

was used to assess biodistribution.

Imaging was performed at days 1, 3, 7, 11, 14, and

17.

MSCs did not distribute systemically. MSCs

tended to migrate from the joint to the place of the

lesion.

J. Clin. Med. 2021, 10, 2925

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Table 4. Cont.

Article

Model

Disease

(Number of

Animals)

Route of

Administration

(Source of Cells)

Cell-Marking Technique

Detection Time and Outcome

Comments

Satué et al.

[75] (2019)

Rats

Patellofemoral

cartilage defect

Intraarticular

(allogenic MSCs)

MSCs expressing heat stable human

placental alkaline phosphatase were

used. Histological and

immuno-histochemical analyses

were performed in joint tissue and

distant organs (heart, spleen,

kidney, liver and lung)

Ex vivo analysis was performed at 1 day, 1 week, 1,

2 and 6 months.

Injected MSCs remained in the synovial cavity,

engrafted within the cartilage lesion, and were

detectable up to 1 month post-injection. No

systemic distribution was observed, apart from 1

case of MSCs in the lung.

Li et al. [67]

(2016)

Mice

Osteoarthritis

Intraarticular

(xenogenic

MSCs—human

MSCs)

MSCs were labeled with DiD

ﬂuorescent dye. In vivo

bioluminescence imaging, and ex

vivo quantitative PCR were

performed to assess biodistribution.

Ex vivo imaging was performed up to day 70. PCR

was performed at day 14 and 70 in heart, liver,

spleen, lung, kidney, brain, muscle adjacent to the

joint, and the whole injected knee join.

MSCs were detected in the injected joint up to day

70 in diseased mice. In healthy mice, MSCs were

detected up to day 21.

No systemic distribution of MSCs was found.

MSCs seem to stand long

times in the injected joint

with no systemic

distribution.

Marquina

et al. [104]

(2017)

Rats

Intraarticular

chondrocyte

trasplantation

Intraarticular,

intravenous,

intraperitoneal

(allogenic MSCs)

MSCs were labeled with luciferase.

Bioluminescence imaging was

performed to assess biodistribution.

Imaging was performed at 2 h, 24 h, 2, 4 and 5 days.

After intraarticular injection, no distribution of

MSCs was detected.

After intravenous injection, most MSCs were

trapped in the lungs and disappeared within 24 h.

After intraperitoneal injection, MSCs were

localized in the injection site without distribution

up to 5 days.

Li et al. [68]

(2017)

Rats

Osteoarthritis

Intraarticular

(xenogenic

MSCs—human

MSCs)

MSCs were labeled with DiD

ﬂuorescent dye. In vivo

bioluminescence imaging and ex

vivo histologic examinations were

performed.

In vivo imaging was performed up to 70 weeks.

MSCs were detected in the injected join up to

9 weeks. No systemic distribution was observed.

MSCs seem to stand long

times in the injected joint

with no systemic

distribution.

J. Clin. Med. 2021, 10, 2925

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Table 4. Cont.

Article

Model

Disease

(Number of

Animals)

Route of

Administration

(Source of Cells)

Cell-Marking Technique

Detection Time and Outcome

Comments

Meseguer-

Olmo et al.

[21] (2017)

Rabbits

Healthy animals

Intraarticular and

intravenous

(xenogenic

MSCs—human

MSCs)

MSCs were labeled

with^99mTc-HMPAO. Scintigraphic

images and qPCR in tissues (liver,

kidney, heart, lung, bladder, knee,

gallbladder) were used for assessing

biodistribution.

Images were taken every 30 s during 25 min. qPCR

was performed at 24 h.

Intravenous MSCs distributed mainly to the lungs.

Intraarticular MSCs did not distributed.

Toupet et al.

[66] (2015)

Mice

Osteoarthritis

and arthritis

(unknown

number)

Intravenous and

intraarticular

(xenogenic

MSCs—human

MSCs)

Human MSCs were infused,

Quantitative assays for human

DNA and mRNA were used to

evaluate the distribution in 13

different organs.

Necropsies were performed at different times (1,

10, 30, 42) and PCR was performed.

After intravenous infusion, MSCs were only

detected in lungs in day 1. No MSCs were detected

in day 10.

After intra-articular injection, MSCs were detected

for at least 10 days in osteo-arthritic knee joints.

No MSCs were detected in other organs after in

these mice.

After intra-articular

injection, MSCs do not seem

to distribute to other organs

or tissues.

Shim et al.

[73] (2015)

Mice

Osteoarthritis

and healthy

models

Intraarticular and

intravenous

(xenogenic

MSCs—human

MSCs)

Human MSCs were injected and

qPCR tests were used to assess

biodistribution in the different

organs.

At 15 min and 8 h after injection, samples were

collected from eight organs (spleen, kidney, liver,

lymph nodes, muscle, lung, heart, brain). Blood

concentrations were also monitored.

After intravenous injection MSCs were detected

immediately in blood, with a progressive decrease.

After intraarticular injection, MSCs were detected

in blood with a peak at 8 h.

No systemic distribution was observed after

intraarterial delivery. After intravenous injection,

most MSCs were trapped in the lungs.

After intraarterial injection,

MSCs are detectable in

blood with a peak at 8 h.

However, no systemic

distribution is observed.

Delling et al.

[69] (2015)

Sheep

Osteoarthritis

Intraarticular

(autogenic MSCs)

MSCs were labelled with SPION

particles.

MRI and histological analyses were

performed.

MR images were acquired at injection and at 1, 4, 8,

and 12 weeks. Ex vivo histological examination

was performed at 12 weeks.

MSCs were found in the joint up to 12 weeks,

without systemic distribution.

J. Clin. Med. 2021, 10, 2925

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Table 4. Cont.

Article

Model

Disease

(Number of

Animals)

Route of

Administration

(Source of Cells)

Cell-Marking Technique

Detection Time and Outcome

Comments

Ikuta et al.

[76] (2015)

Rats

Healthy and

cartilage defect

models

Intraarticular (a

magnet was used for

selective

accumulation of

MSCs) and

intravenous

(xenogenic

MSCs—human

MSCs)

MSCs were labeled with DiR

ﬂuorescent dye and iron

nanoparticles.

MRI and ﬂuorescent imaging were

used to assess biodistribution.

Histological exams were also

performed.

Bioluminescence imaging was performed

immediately and 1, 3, 7, 14, 21, and 28 days after

cell transplantation. At day 28, organs were

collected for ex vivo analyses. After intraarticular

injection, MSCs remained in the joint. The use of

the magnet led to magnetic MSCs accumulation in

the target lesion.

The use of a magnet during

magnetic-labeled MSCs

transplantation can lead to

selective accumulation of

cells into the cartilage

defects.

Table 5. Biodistribution after intralesional administration (except for intra-central nervous system) of MSCs in animal models.

Article

Model

Disease

(Number of

Animals)

Route of

Administration

(Source of Cells)

Cell-Marking Technique

Detection Time and Outcome

Comments

Dave et al.

[54] (2017)

Mice

Chronic bowel

inﬂammation

Intra-cardiac

(xenogenic

MSCs—human

MSCs)

MSCs were labeled with luciferase

and red ﬂuorescent protein.

In vivo and ex vivo

bioluminescence and histologic

examinations were performed to

assess biodistribution.

Images were taken up to 24 h after injection.

Histology was performed at 24 h post injection.

MSCs in healthy mice distributed mainly to

lungs, spleen and liver. In contrast, MSCs in

diseased mice were located mainly in the

intestine, with low pulmonary captation.

After intracardiac injection,

MSCs are able to distribute

mainly to the inﬂamed

intestine.

Jiang et al.

[109] (2018)

Rats

Myocardial

infarction model

(repeated

ischemia model)

Intra-myocardial

(allogenic MSCs)

MSCs were harvested from male

rats and injected into female rats.

qPCR was performed in different

tissues to assess biodistribution

(heart, lungs, spleen and liver)

Examinations were performed 3 weeks after

injection.

MSCs had a greater homing in heart and a

lower distribution to peripheral organs when

repeated ischemia was applied.

J. Clin. Med. 2021, 10, 2925

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Table 5. Cont.

Article

Model

Disease

(Number of

Animals)

Route of

Administration

(Source of Cells)

Cell-Marking Technique

Detection Time and Outcome

Comments

Bansal et al.

[108] (2015)

Mice

Healthy model

Intra-myocardial and

intravenous

(allogenic MSCs and

xenogenic

MSCs—Human

MSCs)

MSCs were labeled with

89Zr-desferrioxamine. PET scans

and radioactivity analyses were

performed to assess biodistribution

PET was performed at days 2, 4, and 7. Ex vivo

radioactivity analyses were performed at day 7.

After intra-myocardial injection, MSCs were

retained in the myocardium, as well as

redistributed to the lung, liver, and bone.

Intravenously administered MSCs also

distributed primarily to the lung, liver, and

bone.

Blazquez et al.

[107] (2015)

Pigs

Myocardial

infarction model

Intrapericardial

(allogenic MSCs)

MSCs were labelled with SPION

particles.

Biodistribution was assessed with

MRI, histology and PCR.

MRI was performed at days 3, 5 and 7.

MSCs were detected to home mainly in the left

ventricle. They were also detected in the right

ventricle, and both atriums.

After intrapericardial

injection, MSCs distribute

mainly to left ventricle.

Lebouvier

et al. [78]

(2015)

Pigs and

mice

Osteonecrosis of

the femoral head

Intraosseous

(xenogenic

MSCs—Human

MSCs)

Human MSCs were injected and

qPCR, cytometry and histologic

analysis was performed to assess

biodistribution in different tissues

(Femoral head, adyacent tissues,

liver, kidneys, spleen, and lungs).

Tissues were collected at either 30 min or 24 h

after injection.

No MSCs were detected in other organs apart

from the injection site.

Khan et al.

[71] (2018)

Mice

Tendon injury

Intralesional

(autogenic MSCs)

MSCs were labelled with

ﬂuorescent-conjugated magnetic

iron-oxide nanoparticles (MIONs)

and were tracked with MRI,

histology and ﬂow cytometry.

Tendons were recovered post mortem at 1 day,

and 1–2, 4, 12 and 24 weeks after MSC injection.

MSCs distributed throughout the tendon

synovial sheath but restricted to the synovial

tissues, with no MSCs detected in the tendon

or surgical lesion. After day 14, no MSCs were

detected.

J. Clin. Med. 2021, 10, 2925

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Table 5. Cont.

Article

Model

Disease

(Number of

Animals)

Route of

Administration

(Source of Cells)

Cell-Marking Technique

Detection Time and Outcome

Comments

Burk et al.

[72] (2016)

Horse

Tendon injury

Intralesional

(autogenic MSCs)

MSCs were 10106 Molday ION

Rhodamine B-labeled.

Biodistribution was assesd with

MRI, ﬂow cytometry and histology

Tracking techniques were performed up to

24 weeks after injection. Labeled cells could be

traced at their injection site by MRI as well as

histology for the whole follow-up period of

24 weeks. Furthermore, small numbers of

labeled cells were identiﬁed in peripheral

blood within the ﬁrst 24 h after cell injection

and could also be found until week 24 within

the contralateral control tendon lesions that

had been injected with serum

Ryska et al.

[106] (2017)

Rats

Fistula model in

Crohn’s disease.

Intralesional

(periﬁstula)

(allogenic)

MSCs were labeled with luciferase.

Bioluminescence imaging was

performed to assess biodistribution.

Imaging was performed at days 0, 2, 7, 14 and

30. MSCs distributed only in the injection site,

with a high reduction of luminescence by day 2.

MSCs were detectable up to day 30.

No systemic distribution

was shown after

intralesional injection.

Zhu et al. [82]

(2015)

Rats

Ovarian injury

Intraovaric and

intravenous

(xenogenic

MSCs—Human

MSCs)

MSCs were ﬂuorescent labeled with

PKH26. Ex vivo bioluminescence

techniques were used to assess

biodistribution (brain, liver, kidney,

urocyst, ovary and uterus were

collected).

Bioluminescence was performed 1, 15, 30 and

45 days after injection.

After intraovaric injection, MSCs were detected

only in ovaries and uterus. After intravenous

injection MSCs were detected in liver, kidney,

ovary and uterus.

Sadeghi et al.

[79] (2016)

Rats

Birth-trauma

injury (urinary

disfunction)

(285 animals)

Intraurethral and

intravenous

(xenogenic

MSCs—Human

MSCs)

Alu genomic repeat staining,

PKH26 labeling, and

luciferase-expression labeling.

Histologic, genetic and

bioluminescence tests were

performed to evaluate MSCs

distribution.

Different assessments were performed at 0, 1, 4

and 10 days after injection.

No positive Alu-stained nuclei were observed

in urethras at 4, 10, and 14 days.

PKH26-labelled cells were found in all urethras

at 2 and 24 h. Bioluminescence study showed

increased luciferase expression from day 0 to 1

following injection, with a progressive

disappearance until day 7.

No MSCs were detected in

periurethral tissue after

intravenous injection.

MSCs were detected for less

than 7 days in periurethral

tissues after local injection.

J. Clin. Med. 2021, 10, 2925

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Table 5. Cont.

Article

Model

Disease

(Number of

Animals)

Route of

Administration

(Source of Cells)

Cell-Marking Technique

Detection Time and Outcome

Comments

Li et al. [136]

(2017)

Rabbits

Chronic

salpingitis model

Intrauterus and

intravenous

(xenogenic

MSCs—Human

MSCs)

MSCs were labeled with green

ﬂuorescent protein and cyto-keratin

7. Ex vivo bioluminescence imaging

was performed in different organs

(oviduct, uterus, liver, and bladder).

The assessment was performed 1 week after

perfusion.

No clear results are derived from this study.

MSCs were detected in the uterus, bladder and

oviduct.

Ryu et al. [80]

(2018)

Sprague-

Dawley

rats

Interstitial

cystitis/bladder

pain sindrome

(unknown

number)

Injection into the

outer layer of the

bladder.

(xenogenic

MSCs—Human

MSCs)

Genetic transduction with green

ﬂuorescent protein was wed for

labelling.

Longitudinal microcystoscopy

(combining confocal microscopy

and cystoscopy) was used to assess

the distribution of MSCs.

Images were obtained between 3 and 42 days

after transplantation.

The number of cells detected decreased rapidly

until day 7 and later decreased gradually until

day 42.

After day 30, MSCs migrated from the serosa

and muscularis layers to the urothelium. At

day 30, most of the cells were distributed in

vascular structures.

MSCs are capable of

migrating through the layers

of the bladder and might be

able to differentiate into

perivascular cells after day

30 post injection.

Dou et al.

[81] (2019)

Rats

Erectile

dysfuncion

(unknown

number)

Intra-cavernosal.

(xenogenic

MSCs—Human

MSCs)

MSCs were labelled with mKATE

and Renilla reniformis luciferase.

Bioluminescence was used to assess

the biodistribution. Histologic

samples were obtained from penis,

kidney, liver, lung, heart, skin,

prostate, testis and spleen.

Bioluminescence was performed immediately

after injection and up to 60 min. Histologic

samples were obtained at days 1, 3 and 7 after

injection.

In vivo, MSCs immediately distributed in the

para-penile region. An early migration to the

abdominal area was noted, where the cells

remained up to day 1.

Histologic examinations showed MSCs in the

penile, kidney, prostate and hepatic tissues.

Bioluminescence might be

less sensitive to detect MSCs

in distant tissues.

J. Clin. Med. 2021, 10, 2925

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Table 5. Cont.

Article

Model

Disease

(Number of

Animals)

Route of

Administration

(Source of Cells)

Cell-Marking Technique

Detection Time and Outcome

Comments

Kallmeyer

et al. [114]

(2020)

Rats

Cutaneous

wound

Intradermal and

intravenous

(allogenic MSCs)

MSCs were labeled with luciferase

and green ﬂuorescent protein.

Bioluminescence imaging and

immunohistological analysis were

performed to assess biodistribution.

Imaging was performed at 3 h, 24 h, 48 h, 72 h

and 7 and 15 days.

Intravenous MSCs were detected in the lungs

3 h after injection with a signal disappearance

from 72 h. No MSCs were detected in the

wound. Locally administered MSCs remained

strongly detectable for 7 days at the injection

site without systemic distribution.

Tappenbeck

et al. [112]

(2019)

Mice

Healty animals

(unknown

number)

Intradermal and

intravenous

(xenogenic

MSCs—Human

MSCs)

Human MSCs were injected and

genetic tests (quantitative PCR)

were done in tissue samples: blood,

skin/subcutis and skeletal muscle

at the injection site, lymph node,

liver, spleen, lungs, brain, femur

bone, and bone marrow, kidneys,

thymus, thyroid/para-thyroid

gland and ovaries or testes) to

evaluate biodistribution.

After intradermal injection, mice were

sacriﬁced at 1 week, 3 months and 4 months.

After intravenous injection, mice were

sacriﬁced.

After intradermal injection, MSCs were

detected in the skin up to 3 months and also in

draining limph nodes after 1 week. No MSCs

were detected in any other tissues.

After intravenous injection, MSCs were

detected mainly in the skin and muscle near to

the injection site and also in the lungs on day 8.

After 1 month, most MSCs were in the lungs.

MSCs were also detected in low quantities in

kidney and thymus after 1 month.

After intradermal injection,

MSCs seem to remain in the

skin and migrate to lymph

node, without signiﬁcant

systemic distribution.

Zhou et al.

[137] (2017)

Mice

Immune deﬁcient

mice

Intradermal (a slice

of cells).

(xenogenic

MSCs—canine

MSCs)

MSCs were labeled with ultrasmall

super-paramagnetic Fe3O4

nanoparticles (USPIO). MRI was

used to assess biodistribution.

MRI was performed at 1 week, 4 weeks and

12 weeks after transplanting the cell sheets.

MSCs were detected up to 12 weeks with

gradual decrease of the captation.

Pratheesh

et al. [113]

(2017)

Rabbits

Cutaneous

wound

Intradermal

(xenogenic

MSCs—goat MSCs)

MSCs were labeled with PKH26.

Fluorescent microscopy was

performed to assess biodistribution

within the wound.

Skin samples were collected from respective

wounds on 3, 7, 10 and 14 days.

MSCs demonstrated a diffuse pattern of

distribution initially and were later

concentrated towards the wound edges and

ﬁnally appeared to be engrafted with the newly

developed skin tissue.

J. Clin. Med. 2021, 10, 2925

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Table 5. Cont.

Article

Model

Disease

(Number of

Animals)

Route of

Administration

(Source of Cells)

Cell-Marking Technique

Detection Time and Outcome

Comments

Léotot et al.

[77] (2015)

Mice

Immunodeﬁcient

mice

MSCs were

pre-loaded into the

bone graft

(xenogenic

MSCs—Human

MSCs)

Human MSCs were used and qPCR

tests were used to assess

biodistribution.

Constructs and organs (liver, spleen, lungs,

heart, and kidneys) were harvested 24 h or

each week between 1 and 7 weeks after

implantation procedures.

No biodistribution of MSCs was detected.

MSCs were detectable in the graft up to 6

weeks.

Lopez-

Santalla et al.

[103] (2017)

Mice

Colitis

Intranodal injection

(inguinal nodes)

(xenogenic

MSCs—Human

MSCs)

MSCs were labeled with luciferase.

Biodistribution was assessed with

bioluminescence imaging.

Bioluminescence imaging was performed 48 h

after injection.

MSCs mainly remained in the injected lymph

nodes or fat surrounding them 48 h after

injection. No signiﬁcant systemic distribution

was found, although the amount of MSCs in

the intestine was relatively high.

After intranodal injection,

most MSCs remained in the

injection site 48 h later.

Packthongsuk

et al. [105]

(2018)

Pigs

Healthy animals

Intraperitoneal

(autogenic MSCs)

MSCs (in this case, isolated from

Wharton’s Jelly) were labeled with

SRY sequences and PKH26-labeled

Ex vivo evaluation was performed

with qPCR and confocal microscopy.

Tissues were collected from the

heart, lung, pancreas, liver, kidney,

omentum, stomach, intestine,

uterine horn, ovary, muscle, and

bone marrow.

Biodistribution was assessed at 6 h, 24 h, and 7,

14 and 21 days after administration.

All tissues were positive for MSCs for 1-day-,

1-week-, 2-week-, and 3-week-old recipients.

MSCs-injected IP

consistently reached tissues

throughout the body. This

result indicates that

intaperitoneal injection

should be considered in

MSCs transplantations.

Hsu et al.

[102] (2017)

Mice

Severe combined

immunodeﬁ-

ciency

Intrahepatic and

intrasplenic

(xenogenic

MSCs—Human

MSCs)

MSCs were labeled with luciferase,

red ﬂuorescent protein and herpes

simplex virus-1 thymidine kinase.

PET, CT, bioluminescence imaging

and histological analyses were

performed to assess biodistribution.

Images and ex vivo analysis were collected for

weeks 1 to 4.

The intrahepatic group showed a conﬁned

signal at the injection site, while the

intrasplenic group displayed a dispersed

distribution at the upper abdominal liver area,

and a more intense signal.

J. Clin. Med. 2021, 10, 2925

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Table 5. Cont.

Article

Model

Disease

(Number of

Animals)

Route of

Administration

(Source of Cells)

Cell-Marking Technique

Detection Time and Outcome

Comments

Liu et al.

[100] (2017)

Mice

Healthy and

NK-activated

mice

(unknown

number)

Intrahepatic

(xenogenic

MSCs—Human

MSCs)

MSCs were labeled with the

Luc2-mKate2 dual-fusion reporter

gene.

Bioluminescence was performed to

assess biodistribution.

Images were collected at multiple time points.

Bioluminescence imaging showed a gradual

decline in the signal in the liver in both groups.

NK-activated group showed a signiﬁcantly

more rapid decrease in the signal.

NK cells seem to have a role

in the elimination of MSCs

transplanted into the liver.

Xie et al. [99]

(2019)

Rats

Acute liver injury

(unknown

number)

Intrahepatic

(xenogenic

MSCs—Human

MSCs)

MSCs were transduced sith

hHNF4α and luciferase2-mKate2

genes.

Bioluminescence imaging was used

to track their biodistribution.

Imaging was performed immediately after

transplantation and until disappearance of

cells.

MSCs were only distributed in the liver. They

were cleared within a short time after

transplantation.

Yaochite et al.

[101] (2015)

Mice

Stretozotocin-

induced diabetes

mellitus

(unknown

number)

Intrapancreatic and

intrasplenic

(allogenic MSCs)

MSCs were labelled with d-luciferin.

Bioluminescence imaging

techniques were used to assess the

biodistribution.

In vivo analysis was performed 0, 1, 3, 5, 8 and

11 days after injection. Ex vivo analysis were

performed 2 days after injection.

Intrasplenic MSCs were retained in the spleen

and distributed to the liver, with a progressive

decrease up to 8 days.

Intrapancreatic MSCs did not distribute to

other organs, and had a progressive decrease

up to 8 days.

Instrasplenic MSCs are

capable of distribute to the

liver.

Intrapancreatic MSCs do not

seem to be able to distribute

to other organs.

Lopez-

Santalla et al.

[103] (2018)

Mice

Colitis

Intraperitoneal

(xenogenic

MSCs—Human

MSCs)

MSCs were labeled with luciferase.

Bioluminescence was used to assess

the biodistribution.

Biodistribution of MSCs was measured in the

main organs (liver, spleen, intestine, lungs,

heart and blood) and lymph nodes (LNs,

inguinal, popliteal, parathymic, parathyroid,

mesenteric, caudal and axillary) 48 h after injection.

Most MSCs distributed to abdominal organs

(liver, spleen and intestine), with few

remaining in lymph nodes, lungs, blood and

heart. Biodistribution did not change

significantly between healthy and diseased mice.

Intraperitoneal injection

seems to lead to abdominal

spreading of MSCs.

J. Clin. Med. 2021, 10, 2925

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Table 5. Cont.

Article

Model

Disease

(Number of

Animals)

Route of

Administration

(Source of Cells)

Cell-Marking Technique

Detection Time and Outcome

Comments

Chen et al.

[110] (2020)

Rats

Broncopulmonary

dysplasia

Intratracheal

(xenogenic

MSCs—Human

MSCs)

MSCs were labeled with Green

Fluorescent Protein and luciferase.

Bioluminescence was used to assess

the biodistribution in the lungs.

Images were taken every 5 s up to 1 min.

MSCs distributed in the lungs without

systemic distribution.

Intratracheal injection lacks

systemic distribution of

MSCs.

Cardenes

et al. [111]

(2019)

Sheep

Acute respiratory

syndrome

Intrabronchial and

intravenous

(xenogenic

MSCs—Human

MSCs)

MSCs were labeled with ¹⁸FDG.

PET-TC was performed to assess

biodistribution.

Images were taken 1 and 5 h after cell

administration.

After intrabronchial administration, MSCs

remained in the injection site at 1 and 5 h

without systemic distribution.

After intravenous injection, MSCs distributed

widely to organs, but with a preference for the

lungs.

Both administration routes

are convenient for treating

acute respiratory syndrome.

Table 6. Biodistribution after intra-central nervous system administration of MSCs in animal models.

Article

Model

Disease

(Number of

Animals)

Route of

Administration

(Source of Cells)

Cell-Marking Technique

Detection Time and Outcome

Comments

Li et al. [98]

(2015)

Different animals and models

(the article is a review)

Intranasal

(different sources)

Different techniques.

Some results are:

•

MSCs reached an intracerebral glioma site within 6 h after i.n. delivery,

with a further signiﬁcant increase in cell numbers within 24 h;

•

Intranasal application of MSCs resulted in the appearance of cells in the

olfactory bulb, brain and spinal cord, and about one-fourth of MSCs

survived for at least 4.5 months in the brain.

Zhang et al.

[138] (2017)

Rats

Spinal cord

injury

Intra-spinal cord

(allogenic MSCs)

MSCs were labeled with

Gd-DTPA-FA and

neuroﬁlament-200. MRI and

histological examinations were

performed to assess biodistribution.

Examinations were performed at day 1, 7, 14

and 28 post delivery.

In the ﬁrst 7 days, transplanted cells were

observed near the injection point. The number

of cells reached a maximum at day 14 and then

gradually distributed along the segmental

injury. No systemic distribution was observed.

J. Clin. Med. 2021, 10, 2925

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Table 6. Cont.

Article

Model

Disease

(Number of

Animals)

Route of

Administration

(Source of Cells)

Cell-Marking Technique

Detection Time and Outcome

Comments

Barberini et al.

[83] (2018)

Horses

Healthy and

myelopathy-

model animals.

(9 animals)

Intrathecal, both

atlanto-occipital (AO)

and lumbo-sacral

(LS) injection.

(allogenic MSCs)

99mTc-HMPAO was used to label

MSCs.

Later evaluation was performed

with a gamma camera and

histologic samples.

Imaging was performed each hour until 5 h

post-infusion.

MSCs administered by AO

injection were found to distribute caudally

through-out the vertebral canal.

MSCs administered by LS injection did not

distribute cranially.

Histologic tests did not show the presence of

MSCs in diseased areas.

LS injection of MSCs does

not seem to be proper to

treat central nervous system

distant lesions.

Quesada et al.

[85] (2019)

Mice

Non-obese

diabetic severe

combined im-

munodeﬁciency

mice

Intrathecal

(xenogenic

MSCs—Human

MSCs)

Human MSCs were used.

Histologic evaluation and qPCR

were performed in different tissues

(Heart, brain, cerebellum, spinal

cord, liver, spleen, lungs, kidneys

and gonads).

Evaluation was performed 24 h and 4 months

after injection.

24 h post-injection, MSCs were detected in the

spinal cord and in 1 heart.

4 months after injection, MSCs were detected

in 3 hearts and in 1 brain.

Kim et al. [84]

(2020)

Rats

Healthy rats

Intrathecal (injected

via L2-L3 space)

(xenogenic

MSCs—Human

MSCs)

Fluorescent dye DiD was used to

label MSCs. Ex vivo

bioluminescence and qPCR of brain,

spine and heart, lung, liver, spleen,

and kidney was used to assess

biodistribution.

Imaging was performed at 0, 6, and 12 h post

injection. MSCs were detected in the spinal

cord at all times. MSCs were found in the brain

only at 12 h. No other organs showed MSCs.

Increasing the Cell Injection dose of MSCs

improved the migration of MSCs to the brain.

MSCs are able to migrate

from spinal cord to the brain.

This migration can be

improved by the increase of

the dose.

Violatto et al.

[97] (2015)

Mice

Amyotrophic

lateral sclerosis

model

Intracerebral (lateral

ventricles) and

intravenous

(xenogenic

MSCs—Human

MSCs)

MSCs were double labeled with

ﬂuorescent nanoparticles and

Hoechst-33258. Bioluminescence

and histologic examinations were

used to assess biodistribution.

In vivo and ex vivo analyses were performed

at 1, 7, 21 days.

By intravenous administration cells were

sequestered by the lungs and rapidly cleared

by the liver. MSCs transplanted in lateral

ventricles remained on the choroid plexus for

the whole duration of the study even if

decreasing in number. Few cells were found in

the spinal cord, and no migration to brain

parenchyma was observed

J. Clin. Med. 2021, 10, 2925

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Table 6. Cont.

Article

Model

Disease

(Number of

Animals)

Route of

Administration

(Source of Cells)

Cell-Marking Technique

Detection Time and Outcome

Comments

Geng et al.

[91] (2015)

Rats

Cerebral

ischemia

Intracerebral

(allogenic MSCs)

A gadolinium-based cell labeling

technique was used.

MRI images were used to assess

biodistribution.

MRI was used to image the cells 1,3, 5 and

7 days after the Gd-MSC injection.

MSCs did not distribute systemically.

Mastro-

Martinez et al.

[90] (2015)

Rats

Traumatic brain

injury

Intracerebral

(allogenic MSCs)

Green ﬂuorescent protein was used

to label cells.

Histological examinations and

immunochemistry were used to

assess biodistribution.

Histologic examination was performed at 24 h

and 21 days after transplantation.

MSC were found in the perilesional area at

24 h, and their number decreased with time

after transplantation. MSC treatment increased

the cell density in the hippocampus and

enhanced neurogenesis in this area.

Park et al.

[96] (2016)

Beagle

dogs

Healthy animals

Intracerebral

(intra-ventricular)

(xenogenic

MSCs—Human

MSCs)

Human MSCs were used.

Immunohistochemical and qPCR

were performed to assess

biodistribution.

Brains were collected 7 days after infusion.

MSCs migrated from ventricles towards the

cortex, being found in the brain parenchyma,

especially along the lateral ventricular walls.

MSCs were also detected in the hippocampus

and the spinal cord.

No systemic distribution of MSCs was

detected.

Xie et al. [87]

(2016)

Rats

Intracerebral

hemorrhage

Intracerebral and

intravenous

(xenogenic

MSCs—Human

MSCs)

A ﬂuorescent dye was used to label

MSCs (CM-DiI). Histologic

evaluation was used to assess

distribution of MSCs.

Histologic examination was performed at 28

days.

After intracerebral injection, MSCs stayed in

the injection place, distributed around the

hemorrhage. A small amount of cells migrated

to the contralateral hemisphere. After

intravenous injection, MSCs were also found in

the cerebral area.

Both intracerebral and

intravenous routes are

appropriate for treating

intracerebral hemorrhage.

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Table 6. Cont.

Article

Model

Disease

(Number of

Animals)

Route of

Administration

(Source of Cells)

Cell-Marking Technique

Detection Time and Outcome

Comments

Duan et al.

[88] (2017)

Rats

Cerebral

ischemia

(54 animals)

Intracerebral

injection (right

striatum)

Green ﬂuorescent protein MSCs

(GFP-MSCs) and SPION labeled.

MRI and histology were used to

assess biodistribution.

Imaging and/or histology were performed

weekly from week 1 to 8 weeks after cells

transplantation.

MSCs were found to remain in the area in a

high quantity in week 1. Later, MSCs number

decreased drastically, being detectable up to

week 8. A small amount of cells migrated to

corpus callosum.

Dong et al.

[95] (2017)

Rats

Brain traumatic

injury

(30 animals)

Intracerebral

injection

(intraventricular)

(allogenic MSCs)

Green ﬂuorescent protein MSCs

(GFP-MSCs).

Imaging techniques and histology

were used to assess biodistribution

in blood vessels.

Techniques were performed at 10, 14, and

17 days. MSCs were found to home in large

arteries (thoracic aorta, abdominal aorta,

common iliac artery) 10, 14, and 17 days after

transplantation.

MSCs seem to distribute

after brain injury when

injected intraventricularly.

Lee et al. [89]

(2017)

Mice

Familial

Alzheimer’s

disease

Intracerebral

injection (Injection

into the hippocampi)

(xenogenic

MSCs—Human

MSCs)

Ferumoxytol was used to label

MSCs.

MRI and histology were used to

assess biodistribution.

Techniques were performed at 1, 7 and 14 days.

MSCs were found to remain in the injection site

up to 14 days after injection.

Wang et al.

[86] (2018)

Sprague

Dawley

rats

Glioma

(unknown

number)

Intracerebral

(MSCs were injected

contralaterally to

glioma)

(allogenic MSCs)

CM-Dil staining was used to label

MSCs, which also contained

Paclitaxel.

Confocal laser-scanning microscopy

was used to assess the distribution

of MSCs. Later histological

examinations assessed the

distribution of MSCs within the

brain.

Necropsies were performed 2 days after MSCs

injection.

MSCs were distributed in clusters in the

injection area, and were also found within the

glioma.

MSCs seem to spread within

a short period of time from

one hemisphere to another,

after intracerebral injection.

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Table 6. Cont.

Article

Model

Disease

(Number of

Animals)

Route of

Administration

(Source of Cells)

Cell-Marking Technique

Detection Time and Outcome

Comments

Mezzanote

et al. [94]

(2017)

Mice

Healthy mice

(unknown

number)

Intracerebral

injection (brain

cortex)

MSCs were transfected with a novel

bioluminescent/near infrared

ﬂuorescent (NIRF) fusion gene.

Fluorescence images and

bioluminescence were used to

assess the distribution of the cells.

Images were taken up to week 7 after

transplantation.

Movement of the MSCs was not assessed.

MSCs were detected for 7 weeks without a

signiﬁcant drop in bioluminescent signals,

suggesting the sustained viability of hMSCs

transplanted into the cortex.

No speciﬁc biodistribution

assessment.

Da Silva et al.

[92] (2019)

Rats

Ischemic stroke

model

Intracerebral

injection

(xenogenic

MSCs—Human

MSCs).

MSCs were labeled with luciferase

and multimodal nanoparticles with

iron. In vivo bioluminescence,

near-infrared imaging and ex vivo

MRI were used to assess

biodistribution.

Biodistribution was assessed at 4 h and 6 days

after cell injection.

MSCs did not distribute. The amount of MSCs

decreased drastically from 4 h to 6 days.

Ohki et al.

[93] (2020)

Rats

Healthy model

Intracerebral

injection

(xenogenic

MSCs—Human

MSCs).

MSCs were labeled with SPIO or

USPIO. MRI and histological

examinations were performed to

assess biodistribution.

MRI images were obtained immediately and at

7- and 14-days post injection.

No MSCs demonstrated migration.

Sukhinich

et al. [53]

(2020)

Rats

Healthy model

Intracerebral and

selective

intra-arterial

(internal carotid

artery)

(xenogenic

MSCs—Human

MSCs).

MSCs were labeled with SPION and

PKH26.

MRI imaging and histology were

performed to assess biodistribution.

The distribution and migration of MSC were

analyzed by MRI from day 1 to day 15, and

histological methods on days 1, 2, 3, 7, and 15.

After intracerebral injection, MSCs moved to

corpus callosum and blood vessels.

After intraarterial injection, most MSCs were

detected in the ipsilateral hemisphere and most

of them within the blood vessels.

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Table 6. Cont.

Article

Model

Disease

(Number of

Animals)

Route of

Administration

(Source of Cells)

Cell-Marking Technique

Detection Time and Outcome

Comments

Teo et al.

[139] (2015)

Mice

Dermal

inﬂammation

(unknown

number)

Retro-orbital

injection.

(xenogenic

MSCs—Human

MSCs).

MSCs were labelled with speciﬁc

techniques for intravital confocal

microscopy (DiI, DiO, DiD or DiR

solution).

Later confocal microscopy was used

to assess the histologic distribution

of MSCs

Imaging was performed 2 h, 4 h and 6 h after

the MSC had been infused. When MSCs were

detected, images were taken every 5 min.

By 2 h post-infusion, arrested and

transmigrating MSC were equally distributed

within both small capillaries and larger

venules. These MSCs were in contact with

neutrophil-platelet clusters.

Platelet depletion led to signiﬁcantly reduced

the preferential homing of MSC to the inﬂamed

Authors concluded that

MSCs transmigrate to

tissues due to the existence

of an active adhesion

mechanism.

Platelets seem to play a

crucial role in MSCs

trafﬁcking.

Table 7. Articles regarding biodistribution of MSCs in humans.

Article

Disease (Number

of Patients)

Route of

Administration

(Source of Mscs)

Cell-Marking Technique

Detection Time and Outcome

Comments

Krueger et al.

[126] (2018)

Breast cancer (28

patients) [116]

Intravenous

(autogenic MSCs)

Flow citometry

MSCs were detected for several hours post-infusion in

peripheral blood.

MSCs are rapidly (less than 1 h) cleared from peripheral

blood after intravenous infusion

The presence of MSCs in

peripheral blood was not

detected after 1 h

post-infusion.

Cirrhosis

(4 patients) [115]

Intravenous

(autogenic MSCs)

111In-oxine labeled

mesenchymal stem cells,

evaluated with Dual head

gamma camera and SPECT

imaging techniques.

MSCs were detected at 2 h, 4 h, 6 h, 24 h, 48 h, 7th and

10th days after infusion.

Pre-48 h images showed a large majority of cells

distributed in the lungs. Later images showed a drastic

decrease in lung captation, with a higher amount of

MSCs distributed in the spleen and few in liver.

There was a clear initial

biodistribution in lungs,

which decreased after 48 h.

Spleen captation was higher

than liver captation, maybe

due to splenomegaly.

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Table 7. Cont.

Article

Disease (Number

of Patients)

Route of

Administration

(Source of Mscs)

Cell-Marking Technique

Detection Time and Outcome

Comments

Henriksson et al.

[118] (2019)

Intervertebral disc

degeneration

(4 patients)

Intralesional

(autogenic MSCs)

MSCs were labeled with iron

sucrose (Venofer^®).

Histologic examinations were

performed to detect the cells.

Intravertebral discs were explanted at 8 months

(3 patients) and 28 months (1 patient) post injection.

MSCs were detected at 8 months, but not at 28 months.

Detected MSCs had differentiated into chondrocyte-like

cells.

MSCs seem to home in

intravertebral discs after

intralesional injection for

long periods of time.

Sokal et al. [117]

(2017)

Haemophilia A

(1 patient)

Intravenous

(Adult-derived human

liver stem cells)

(allogenic MSCs)

MSCs were labeled with

111In-Oxine and

biodistribution was assessed

with sequential planar

imaging (SPECT, TC).

Total body imaging was performed at 1, 4, 24, 48, 72, and

144 h postinfusion.

MSCs were initially (1 h) trapped in the lungs and liver.

At 24 h, MSCs were detected in the right ankle (where

hemarthrosis was recurrent). Up to day 6, lungs signal

decreased and liver signal increased. MSCs were also

detected in small amounts in spleen and bone marrow.

MSCs infusion seemed to

result in an improved

bleeding phenotype and was

well tolerated. Moreover, the

distribution of MSCs to the

place of bleeding suggests

possible in situ production of

factor VIII.

Sood et al. [122]

(2015)

Type 2 diabetes

mellitus

(21 patients)

Intravenous and

selective intraarterial

(superior

pancreaticoduodenal

artery and proximal

splenic artery)

(autogenic bone marrow

MSCs).

MSCs were labeled with

18-FDG.

PET-TC images were used to

assess biodistribution

Images were taken at 30 and 90 min post infusion.

In the intravenous group, MSCs distributed to lungs at

30 min with signiﬁcant clearance in the delayed 90-min

image, with no distribution to pancreas. Selective

intraarterial delivery led to MSCs homing in pancreas

head (pancreaticoduodenal artery) or body (splenic

artery).

Selective intraarterial

delivery leads to selective

homing of MSCs into the

pancreas.

Lezaic et al. [119]

(2016)

Idiopathic dilated

cardiomyopathy

(35 patients)

Intracoronary infusion

(autogenic MSCs)

MSCs were labeled with

99mTc-HMPAO.

Gamma-cammera images

were taken to assess

biodistribution.

Imaging was performed 1 h and 18 h after transplantation.

At 1 h after intracoronary administration, the majority of

MSCs accumulated in the liver, spleen and bone marrow.

Accumulation of MSCs in the myocardium ranged from 0

to 1.45% of injected activity in the ﬁeld of view. The

distribution of labeled stem cells in the myocardium

corresponded to the area supplied by the vessel used for

administration. At imaging 18 h post injection, the

distribution of labeled stem cells appeared unchanged,

but with decreased activity.

The retention of MSCs in the

myocardium is low after

intracoronary injection.

J. Clin. Med. 2021, 10, 2925

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Author Contributions: Conceptualization M.S.-D., S.A.-S. and A.M.-L. Search strategy: M.S.-D.,

M.I.Q.-V., R.S.d.l.T. and T.M.-V. Article review M.S.-D., T.M.-V. and A.S.-S. Expert revision: A.M.-L.

and S.A.-S. Manuscript preparation M.S.-D., T.M.-V., A.S.-S., M.I.Q.-V. and R.S.d.l.T. All authors have

read and agreed to the published version of the manuscript.

Funding: This research has received competitive funding in the call for grants for the ﬁnancing of

Research, Development and Innovation in Biomedicine and Health Sciences in Andalusia, for the

year 2019 (PIGE-0247-2019; PIGE-0242-2019).

Institutional Review Board Statement: Not applicable because no humans or animals were present

in a systematic review.

Informed Consent Statement: Not applicable.

Data Availability Statement: Not applicable.

Conﬂicts of Interest: The authors declare no conﬂict of interest.

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